<!DOCTYPE article PUBLIC "-//NLM//DTD Journal Archiving and Interchange DTD v2.3 20070202//EN" "http://dtd.nlm.nih.gov/publishing/2.3/journalpublishing.dtd">
<article article-type="review-article" xmlns:xlink="http://www.w3.org/1999/xlink">
   <front>
      <journal-meta>
         <journal-id journal-id-type="nlm-ta">Diagn Pathol</journal-id>
         <journal-title>Diagnostic Pathology</journal-title>
         <issn pub-type="epub">1746-1596</issn>
         <publisher>
            <publisher-name>BioMed Central</publisher-name>
            <publisher-loc>London</publisher-loc>
         </publisher>
      </journal-meta>
      <article-meta>
         <article-id pub-id-type="publisher-id">1746-1596-1-32</article-id>
         <article-id pub-id-type="pmid">16999866</article-id>
         <article-id pub-id-type="doi">10.1186/1746-1596-1-32</article-id>
         <article-categories>
            <subj-group subj-group-type="heading">
               <subject>Review</subject>
            </subj-group>
         </article-categories>
         <title-group>
            <article-title>Prevention of an additional surgery for regional lymphadenectomy in melanoma: rapid intraoperative immunostaining of sentinel lymph node imprint smears</article-title>
         </title-group>
         <contrib-group>
            <contrib contrib-type="author" corresp="yes" id="A1">
               <name>
                  <surname>Shidham</surname>
                  <given-names>Vinod B</given-names>
               </name>
               <xref ref-type="aff" rid="I1">1</xref>
               <email>vshidham@mcw.edu</email>
            </contrib>
            <contrib contrib-type="author" id="A2">
               <name>
                  <surname>Komorowski</surname>
                  <given-names>Richard</given-names>
               </name>
               <xref ref-type="aff" rid="I1">1</xref>
               <email>rkomor@mcw.edu</email>
            </contrib>
            <contrib contrib-type="author" id="A3">
               <name>
                  <surname>Neuberg</surname>
                  <given-names>Marcelle</given-names>
               </name>
               <xref ref-type="aff" rid="I2">2</xref>
               <email>mneuburg@mcw.edu</email>
            </contrib>
            <contrib contrib-type="author" id="A4">
               <name>
                  <surname>Walker</surname>
                  <given-names>Alonzo</given-names>
               </name>
               <xref ref-type="aff" rid="I3">3</xref>
               <email>awalker@mcw.edu</email>
            </contrib>
            <contrib contrib-type="author" id="A5">
               <name>
                  <surname>Campbell</surname>
                  <given-names>Bruce H</given-names>
               </name>
               <xref ref-type="aff" rid="I4">4</xref>
               <email>bcampbel@mcw.edu</email>
            </contrib>
            <contrib contrib-type="author" id="A6">
               <name>
                  <surname>Chang</surname>
                  <given-names>Chung-Che</given-names>
               </name>
               <xref ref-type="aff" rid="I5">5</xref>
               <email>jeffchang@pol.net</email>
            </contrib>
            <contrib contrib-type="author" id="A7">
               <name>
                  <surname>Dzwierzynski</surname>
                  <given-names>William W</given-names>
               </name>
               <xref ref-type="aff" rid="I6">6</xref>
               <email>billd@mcw.edu</email>
            </contrib>
         </contrib-group>
         <aff id="I1"><label>1</label>Department of Pathology, Medical College of Wisconsin, Milwaukee, WI, USA</aff>
         <aff id="I2"><label>2</label>Department of Dermatology, Medical College of Wisconsin, Milwaukee, WI, USA</aff>
         <aff id="I3"><label>3</label>Department of Otolaryngology, Medical College of Wisconsin, Milwaukee, WI, USA</aff>
         <aff id="I4"><label>4</label>Department of Surgery, Medical College of Wisconsin, Milwaukee, WI, USA</aff>
         <aff id="I5"><label>5</label>Department of Pathology, The Methodist Hospital, Houston, TX, USA</aff>
         <aff id="I6"><label>6</label>Department of Plastic Surgery, Medical College of Wisconsin, Milwaukee, WI, USA</aff>
         <pub-date pub-type="collection">
            <year>2006</year>
         </pub-date>
         <pub-date pub-type="epub">
            <day>25</day>
            <month>09</month>
            <year>2006</year>
         </pub-date>
         <volume>1</volume>
         <fpage>32</fpage>
         <lpage>32</lpage>
         <ext-link ext-link-type="uri" xlink:href="http://www.diagnosticpathology.org/content/1/1/32"/>
         <history>
            <date date-type="received">
               <day>13</day>
               <month>9</month>
               <year>2006</year>
            </date>
            <date date-type="accepted">
               <day>25</day>
               <month>9</month>
               <year>2006</year>
            </date>
         </history>
         <permissions>
            <copyright-statement>Copyright &#x000a9; 2006 Shidham et al; licensee BioMed Central Ltd.</copyright-statement>
            <copyright-year>2006</copyright-year>
            <copyright-holder>Shidham et al; licensee BioMed Central Ltd.</copyright-holder>
            <license license-type="open-access" xlink:href="http://creativecommons.org/licenses/by/2.0">
               <p>This is an Open Access article distributed under the terms of the Creative Commons Attribution License (<ext-link ext-link-type="uri"
                     xlink:href="http://creativecommons.org/licenses/by/2.0"/>), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
                  cited.</p>
               <!--<rdf xmlns="http://web.resource.org/cc/" xmlns:rdf="http://www.w3.org/1999/02/22-rdf-syntax-ns#" xmlns:dc="http://purl.org/dc/elements/1.1" xmlns:dcterms="http://purl.org/dc/terms"><Work xmlns:dc="http://purl.org/dc/elements/1.1/" xmlns:dcterms="http://purl.org/dc/terms/" rdf:about=""><license rdf:resource="http://creativecommons.org/licenses/by/2.0"/><dc:type rdf:resource="http://purl.org/dc/dcmitype/Text"/><dc:author>
               Shidham
               B
               Vinod
               
               vshidham@mcw.edu
            </dc:author><dc:title>
            Prevention of an additional surgery for regional lymphadenectomy in melanoma: rapid intraoperative immunostaining of sentinel lymph node imprint smears
         </dc:title><dc:date>2006</dc:date><dcterms:bibliographicCitation>Diagnostic Pathology 1(1): 32-. (2006)</dcterms:bibliographicCitation><dc:identifier type="sici">1746-1596(2006)1:1&#x0003c;32&#x0003e;</dc:identifier><dcterms:isPartOf>urn:ISSN:1746-1596</dcterms:isPartOf><License rdf:about="http://creativecommons.org/licenses/by/2.0"><permits rdf:resource="http://web.resource.org/cc/Reproduction" xmlns=""/><permits rdf:resource="http://web.resource.org/cc/Distribution" xmlns=""/><requires rdf:resource="http://web.resource.org/cc/Notice" xmlns=""/><requires rdf:resource="http://web.resource.org/cc/Attribution" xmlns=""/><permits rdf:resource="http://web.resource.org/cc/DerivativeWorks" xmlns=""/></License></Work></rdf>-->
            </license>
         </permissions>
         <abstract>
            <sec id="id1">
               <title>Background</title>
               <p>Sentinel lymph node (SLN) biopsy is performed at many institutions and is considered a standard of care in the management of cutaneous melanoma. The discriminatory immunostaining
                  pattern with the 'MCW Melanoma Cocktail' (a mixture of MART-1 {1:500}, Melan- A {1:100}, and Tyrosinase {1:50} monoclonal antibodies) allows intraoperative immunocytochemical
                  evaluation of imprint smears of SLNs for melanoma metastases. Cohesive cells of benign capsular melanocytic nevi that were also immunoreactive with the cocktail do not exfoliate
                  easily for imprint smear detection.</p>
            </sec>
            <sec id="id2" sec-type="methods">
               <title>Methods</title>
               <p>We prospectively evaluated 73 lymph nodes (70 SLN &#x00026; 3 non-SLN) from 41 cases (mean 1.8, 1 to 4 SLNs/case) of cutaneous melanoma using a rapid 17-minute immunostaining
                  previously published protocol. The results were compared with permanent sections also immunostained with 'the cocktail'.</p>
            </sec>
            <sec id="id3">
               <title>Results</title>
               <p>19.5%, 8/41 cases (12%, 9/73 lymph nodes) were positive for melanoma metastases on permanent sections immunostained with the 'MCW melanoma cocktail'. Melanoma metastases in 87.5%
                  (7/8) of these cases were also detected in rapidly immunostained imprint smears, with 100% specificity and 90% sensitivity. None of the 7 SLNs from 7 cases with capsular nevi showed
                  false positive results.</p>
            </sec>
            <sec id="id4">
               <title>Conclusion</title>
               <p>Melanoma metastases could be detected in imprint smears immunostained with 'MCW Melanoma Cocktail' utilizing a rapid intraoperative protocol. The cohesive cells of the capsular nevi
                  do not readily exfoliate and do not lead to false positive interpretation. In a majority of positive cases, a regional lymphadenectomy could have been completed during the same
                  surgery for SLN biopsy and wide excision of primary melanoma site, potentially eliminating the need for an additional surgery.</p>
            </sec>
         </abstract>
      </article-meta>
   </front>
   <body>
      <sec id="id5">
         <title>Background</title>
         <p>Skin melanoma is the sixth most common cancer in US with an increasing rate in men [<xref ref-type="bibr" rid="B1">1</xref>]. Due to rapidly progressive nature of the disease, aggressive
            definitive therapy at earliest stage is the best option. Because of associated high morbidity and the scarcity of follow up studies evaluating its survival benefits, the role of routine
            performance of regional lymphadenectomy in all cases irrespective of their status is controversial [<xref ref-type="bibr" rid="B2">2</xref>,<xref ref-type="bibr" rid="B3">3</xref>]. An
            international multi-centre randomized prospective trial by the Intergroup Melanoma Surgical Program, reported that regional node dissection offers increased survival in subset of patients
            with nodal metastases (60 years of age or younger with intermediate-thickness nonulcerative melanomas) [<xref ref-type="bibr" rid="B4">4</xref>]. Sentinel lymph node (SLN) biopsy is
            recommended as a tool to identify this subset of patients, who could then undergo elective regional lymph node dissection [<xref ref-type="bibr" rid="B5">5</xref>]. The negative status of
            SLN for melanoma metastases correlates closely with the negative status of regional lymph nodes [<xref ref-type="bibr" rid="B6">6</xref>].</p>
         <p>Thus, prevailing evidence supports the evaluation of SLN for melanoma metastases as the standard of care to identify subset of patients with metastases in cutaneous melanoma. It is
            practiced throughout the academic centers as well as regional and community hospitals with increasing frequency [<xref ref-type="bibr" rid="B2">2</xref>-<xref ref-type="bibr" rid="B22"
            >22</xref>].</p>
         <p>Although, it is beneficial to complete a regional lymphadenectomy during the same initial surgical procedure for 'SLN biopsy with wide excision of primary melanoma', currently the patient
            has to undergo regional lymphadenectomy at later date under separate anesthesia, if SLNs are positive for melanoma metastases. In the current situation the results can not be obtained
            intraoperatively. Preoperative or intraoperative evaluation technique would result in a cost saving with numerous other benefits [<xref ref-type="bibr" rid="B23">23</xref>-<xref
               ref-type="bibr" rid="B31">31</xref>].</p>
         <p>Frozen-section examination (with or without immunohistochemical evaluation) and the morphological evaluation of imprint cytology smears are some of the methods evaluated for the
            intraoperative examination of SLN in cutaneous melanoma. However, the studies evaluating these methods did not demonstrate encouraging results [<xref ref-type="bibr" rid="B25"
               >25</xref>-<xref ref-type="bibr" rid="B31">31</xref>], primarily due to lack of adequate sensitivity.</p>
         <p>The 'MCW Melanoma Cocktail' (a mixture of MART-1 {1:500}, Melan- A {1:100}, and Tyrosinase {1:50} monoclonal antibodies) demonstrated an excellent discriminatory immunostaining pattern
               [<xref ref-type="bibr" rid="B32">32</xref>,<xref ref-type="bibr" rid="B33">33</xref>]. This has facilitated rapid intraoperative evaluation of SLNs for melanoma micrometastases by
            examining imprint smears of SLNs immunostained with the cocktail [<xref ref-type="bibr" rid="B34">34</xref>,<xref ref-type="bibr" rid="B35">35</xref>]. This was not previously possible
            with conventional immunomarkers such as the S-100 protein and HMB45 due to significant interference caused by non-melanoma cells such as dendritic cells and mast cells resulting in a high
            noise to signal ratio [<xref ref-type="bibr" rid="B33">33</xref>,<xref ref-type="bibr" rid="B36">36</xref>].</p>
         <p>In this article we report the results on a series of cutaneous melanoma cases studied with rapid intraoperative evaluation of SLN for melanoma metastases employing imprint smears
            immunostained with 'MCW melanoma cocktail'.</p>
      </sec>
      <sec id="id6" sec-type="materials|methods">
         <title>Materials and methods</title>
         <sec id="id7">
            <title>Patients</title>
            <p>73 lymph nodes (70 SLNs &#x00026; 3 non-SLNs) from 41 cases with clinically localized cutaneous melanoma were studied prospectively under the IRB approved protocol. The demographics
               are shown in table <xref ref-type="table" rid="T1">1</xref>. As this was as a research study, intraoperative decisions for the continuation of a regional lymphadenectomy were not
               executed regardless of positive results with immunostained imprint smear(s) of SLNs. This was explained to the patients participating in the study as a component of the informed
               consent.</p>
            <table-wrap id="T1" position="float">
               <label>Table 1</label>
               <caption>
                  <p>Demographics of all the cases.</p>
               </caption>
               <table frame="hsides" rules="groups">
                  <thead>
                     <tr>
                        <td/>
                        <td align="left">
                           <bold>Feature</bold>
                        </td>
                        <td align="left">
                           <bold>Details</bold>
                        </td>
                     </tr>
                  </thead>
                  <tbody>
                     <tr>
                        <td align="left">
                           <bold>1</bold>
                        </td>
                        <td align="left">Number of case studied</td>
                        <td align="left">Total- 41 case</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">&#x02003;Males- 21, Females- 20</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">&#x02003;Mean age- 56 years (14 &#x02013; 48 years)</td>
                     </tr>
                     <tr>
                        <td colspan="3">
                           <hr/>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">
                           <bold>2</bold>
                        </td>
                        <td align="left">Total lymph nodes studied</td>
                        <td align="left">73 lymph nodes (SLN- 70, non-SLN- 3*) Mean- 1.7 SLN per case (1&#x02013;4 SLN per case)</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">&#x02003;1 SLN- 16 cases</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">&#x02003;2 SLN- 17 cases</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">&#x02003;3 SLN- 4 cases</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">&#x02003;4 SLN- 4 cases</td>
                     </tr>
                     <tr>
                        <td colspan="3">
                           <hr/>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">
                           <bold>3</bold>
                        </td>
                        <td align="left">Pigmented lesions as indication for SLN biopsy</td>
                        <td align="left">Melanomas- 39</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">Atypical Spitzoid nevus- 1</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">Dysplastic nevus- 1</td>
                     </tr>
                     <tr>
                        <td colspan="3">
                           <hr/>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">
                           <bold>4</bold>
                        </td>
                        <td align="left">Location of primary pigmented lesion</td>
                        <td align="left">Head and neck (15), the trunk (11), the upper extremity (6), and the lower extremity (9).</td>
                     </tr>
                     <tr>
                        <td colspan="3">
                           <hr/>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">
                           <bold>5</bold>
                        </td>
                        <td align="left">Histological type of melanomas (known in 17 cases)<sup>&#x000b6;</sup></td>
                        <td align="left">Superficial spreading- 11</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">Nodular- 5</td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">Superficial spreading with nodular- 1</td>
                     </tr>
                     <tr>
                        <td colspan="3">
                           <hr/>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">
                           <bold>6</bold>
                        </td>
                        <td align="left">Breslow thickness of primary melanoma (known in 38 patients)</td>
                        <td align="left">Mean- 2.1 mm (range 0.7 to 8.9 mm)</td>
                     </tr>
                  </tbody>
               </table>
               <table-wrap-foot>
                  <p>*Case no. 23 (Table 2) had 5 lymph nodes (2 SLN and 3 non-SLN).</p>
                  <p><sup>&#x000b6; </sup>In most cases, the biopsy of primary lesion was performed and interpreted at outside institutions.</p>
               </table-wrap-foot>
            </table-wrap>
            <p>All patients underwent mappings and biopsies of SLNs with a wide excision of the primary site at Froedert Memorial Lutheran Hospital/Medical College of Wisconsin, Milwaukee, WI. A
               standard surgical protocol was used to identify the SLN [<xref ref-type="bibr" rid="B37">37</xref>]. The tumor bed was injected preoperatively with technetium sulfur colloid.
               Additionally, dermis around the lesion was injected intraoperatively with isosulfan blue dye to facilitate the visual identification of the SLN [<xref ref-type="bibr" rid="B37"
               >37</xref>]. The SLNs were harvested and submitted fresh without any fixative to pathology for intraoperative and permanent section evaluation.</p>
         </sec>
         <sec id="id8">
            <title>Pathologic examination (Figure <xref ref-type="fig" rid="F1">1</xref>)</title>
            <fig id="F1" position="float">
               <label>Figure 1</label>
               <caption>
                  <p>Pathological evaluation of sentinel lymph nodes for melanoma metastases. Section number 2, 5, &#x00026; 8- stained with HE; 4- immunostained with 'MCW melanoma cocktail'; 6-
                     negative control; 1, 3, 7, &#x00026; 9- unstained sections on coated slides for immunostaining (but may be used for any stain). Number of slices of SLN shown (a,b,c) is for
                     illustration only and would vary according to the size of the lymph node. (FPTS, formalin-fixed paraffin-embedded tissue sections; H&#x00026;E, hematoxylin and eosin stain;
                     immuno, immunohistochemistry)</p>
               </caption>
               <graphic xlink:href="1746-1596-1-32-1"/>
            </fig>
            <p>The excised lymph nodes were transected perpendicular to the long axis and processed to make imprint smears as previously reported [<xref ref-type="bibr" rid="B35">35</xref>]. The
               imprint smears were immunostained with the 'MCW melanoma cocktail' using either a manual method or the Dako<sup>&#x000ae; </sup>Autostainer with a rapid 17-minute immunostaining
               protocol utilizing the Dako EnvisionTM+ System which does not contain avidin-biotin [<xref ref-type="bibr" rid="B35">35</xref>]. One smear of a previously prepared positive control from
               a known melanoma tumor and processed similar to test smears [<xref ref-type="bibr" rid="B35">35</xref>] was also immunostained with each staining batch.</p>
            <p>The immunostained imprint smears were examined and interpreted as positive, indeterminate, or negative for melanoma metastases. The time required to go from preparation of imprint
               smears to evaluation of immunostained smears was also tracked. Based on the algorithm for intraoperative decision, indeterminate interpretations with immunostained imprint smears were
               considered as negative for statistical analysis.</p>
            <p>After imprint smear preparation, the slices of SLNs were fixed in 10% formalin and processed in the usual manner for paraffin embedding. Three FPTS levels, each cut at
               200-&#x003bc;m intervals were stained with hematoxylin and eosin (H&#x00026;E). The levels adjacent to the middle H&#x00026;E section were immunostained with 'MCW melanoma
               cocktail' using the avidin-biotin-peroxidase complex (ABC) method described previously [<xref ref-type="bibr" rid="B33">33</xref>].</p>
            <p>The results were considered positive for melanoma metastases if cytoplasmic immunoreactivity was detected in cell clusters or individual cells that demonstrated morphologic features of
               melanoma tumor cells. When discrepancies existed between immunostained imprint smears and the immunostained FPTS, the slides were reviewed by another pathologist in an attempt to
               determine the cause of the discrepancy and to arrive at a conclusion (Table <xref ref-type="table" rid="T2">2</xref>).</p>
            <table-wrap id="T2" position="float">
               <label>Table 2</label>
               <caption>
                  <p>Results with rapid intraoperative immunocytochemical evaluation of imprint smears.</p>
               </caption>
               <table frame="hsides" rules="groups">
                  <thead>
                     <tr>
                        <td align="left">
                           <bold>Serial No.</bold>
                        </td>
                        <td align="left">
                           <bold>No. of SLN per case</bold>
                        </td>
                        <td align="left" colspan="3">
                           <bold>SLN</bold>
                        </td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td colspan="3">
                           <hr/>
                        </td>
                     </tr>
                     <tr>
                        <td/>
                        <td/>
                        <td align="left">
                           <bold>IC</bold>
                        </td>
                        <td align="left">
                           <bold>SP</bold>
                        </td>
                        <td align="left">
                           <bold>FR</bold>
                           <sup>4</sup>
                        </td>
                     </tr>
                  </thead>
                  <tbody>
                     <tr>
                        <td align="left">1</td>
                        <td align="left">2</td>
                        <td align="left">B-,C-</td>
                        <td align="left">B-<bold>cn</bold>,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">2</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-<bold>cn</bold></td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">3<sup>&#x000b6;</sup></td>
                        <td align="left">2</td>
                        <td align="left">A1+,A2+,B-</td>
                        <td align="left">A1+,A2-,B-</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">4</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-<bold>cn</bold></td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">5</td>
                        <td align="left">3</td>
                        <td align="left">A-,B-,C-</td>
                        <td align="left">A-,B-<bold>cn</bold>,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">6</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">7</td>
                        <td align="left">2</td>
                        <td align="left">B-,C-</td>
                        <td align="left">B-,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">8</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">9</td>
                        <td align="left">2</td>
                        <td align="left">A-, C-</td>
                        <td align="left">A-, C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">10</td>
                        <td align="left">2</td>
                        <td align="left">A-,C-</td>
                        <td align="left">A-<bold>cn</bold>,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">11*</td>
                        <td align="left">4</td>
                        <td align="left"><bold>A-</bold>,B-,C-,D-</td>
                        <td align="left">A+,B-,C-,D-</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">12<sup>&#x000b6;</sup></td>
                        <td align="left">4</td>
                        <td align="left">A-,B+,C+,<bold>D+</bold></td>
                        <td align="left">A-,B+,C+,D-</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">13<sup>&#x02020;</sup></td>
                        <td align="left">1</td>
                        <td align="left">
                           <bold>A(Intm)</bold>
                        </td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">14</td>
                        <td align="left">1</td>
                        <td align="left">B+</td>
                        <td align="left">B+</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">15</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">16</td>
                        <td align="left">3</td>
                        <td align="left">A+,B-,C-</td>
                        <td align="left">A+,B-,C-</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">17<sup>&#x02020;</sup></td>
                        <td align="left">2</td>
                        <td align="left">A<bold>(Intm)</bold>,B-</td>
                        <td align="left">A-,B-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">18</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">19</td>
                        <td align="left">2</td>
                        <td align="left">A-,B-</td>
                        <td align="left">A-,B-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">20</td>
                        <td align="left">2</td>
                        <td align="left">A1-, C+</td>
                        <td align="left">A1-, C+</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">21</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">22</td>
                        <td align="left">2</td>
                        <td align="left">A-,B-</td>
                        <td align="left">A-,B-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">23<sup>1</sup></td>
                        <td align="left"><sup>1</sup>2</td>
                        <td align="left"><sup>1</sup>A1-,<sup>1</sup>B+,<sup>1</sup>C-,D-,E-</td>
                        <td align="left"><sup>1</sup>A1-,<sup>1</sup>B+,<sup>1</sup>C-,D-,E-</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                     <tr>
                        <td align="left">24</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">25</td>
                        <td align="left">2</td>
                        <td align="left">A-,B-</td>
                        <td align="left">A-,B-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">26</td>
                        <td align="left">2</td>
                        <td align="left">A-,C-</td>
                        <td align="left">A-,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">27</td>
                        <td align="left">1</td>
                        <td align="left">B-</td>
                        <td align="left">B-<bold>cn</bold></td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">28</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">29</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-<bold>cn</bold></td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">30</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">31<sup>2</sup></td>
                        <td align="left">4</td>
                        <td align="left">A-,B-,C-,D-</td>
                        <td align="left">A-,B-,C-,D-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">32</td>
                        <td align="left">2</td>
                        <td align="left">A-,B-</td>
                        <td align="left">A-,B-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">33</td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">34</td>
                        <td align="left">4</td>
                        <td align="left">A-,B-,C-,D-</td>
                        <td align="left">A-,B-,C-,D-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">35</td>
                        <td align="left">3</td>
                        <td align="left">A-,B-,C-</td>
                        <td align="left">A-,B-,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">36<sup>3</sup></td>
                        <td align="left">1</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">37</td>
                        <td align="left">2</td>
                        <td align="left">A-</td>
                        <td align="left">A-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">38</td>
                        <td align="left">2</td>
                        <td align="left">B-,C-</td>
                        <td align="left">B-,C-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">39</td>
                        <td align="left">2</td>
                        <td align="left">A-,B-</td>
                        <td align="left">A-,B-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">40</td>
                        <td align="left">3</td>
                        <td align="left">A-,B-,D-</td>
                        <td align="left">A-,B-,D-</td>
                        <td align="left">N</td>
                     </tr>
                     <tr>
                        <td align="left">41</td>
                        <td align="left">2</td>
                        <td align="left">A-,B+</td>
                        <td align="left">A-,B+</td>
                        <td align="left">
                           <bold>P</bold>
                        </td>
                     </tr>
                  </tbody>
               </table>
               <table-wrap-foot>
                  <p>*One false negative</p>
                  <p><sup>&#x02020;</sup>Two negative cases showed indeterminate results with IC</p>
                  <p><sup>&#x000b6;</sup>Two SLN were positive unequivocally with IC but negative by SP</p>
                  <p><sup>1</sup>Only D&#x00026;E were submitted as sentinel LN (A,B,C were submitted as non-SLN);</p>
                  <p><sup>2</sup>Atypical Spitzoid nevus, recommended SLN biopsy;</p>
                  <p><sup>3</sup>Dysplastic nevus, recommended SLN biopsy;</p>
                  <p><sup>4</sup>Final results summarizing findings in all SLN in a particular case after immunostaining of permanent sections: total number of positive cases- 8 and negative cases-
                     33.</p>
                  <p><italic>Abbreviations in alphabetical order</italic>: +, positive for melanoma metastases; -, negative for melanoma metastases; <bold>cn</bold>, capsular nevus; FR, final result;
                     IC, rapid intraoperative immunocytochemistry; <bold>Intm</bold>, indeterminate for melanoma metastases; LN, sentinel and non-sentinel lymph nodes; N, negative; <bold>P</bold>,
                     positive; SP, final result on permanent sections.</p>
               </table-wrap-foot>
            </table-wrap>
         </sec>
      </sec>
      <sec id="id9">
         <title>Results</title>
         <p>The time required for completing the entire procedure from preparation of imprint smears to evaluation of immunostained smears ranged from 24 to 45 (mean, 35) minutes and depended on the
            size-number of SLNs.</p>
         <p>At the case level, melanoma metastases were identified in lymph nodes of 19.5% (8 out of 41) of patients by the immunohistochemical evaluation of FPTS. 87.5% (7 out of 8) of these cases
            showed metastases in imprint smears immunostained rapidly with the 'MCW melanoma cocktail', potentially saving a second surgery in all these cases. The test demonstrated a sensitivity of
            89%, a specificity of 100%, the negative predictive value of 97%, and the positive predictive value of 100%.</p>
         <p>At the lymph node level, 12% (9/73) nodes were positive for melanoma metastases with immunostained FPTS. Of these, 89% (8/9) showed melanoma metastases in intraoperatively immunostained
            imprint smears utilizing the cocktail demonstrating a sensitivity of 90%, a specificity of 100%, the negative predictive value of 99%, and the positive predictive value of 100%).</p>
         <p>The immunostained tumor cells of melanoma metastases (Figure <xref ref-type="fig" rid="F2">2</xref>) showed a high nuclear to cytoplasmic ratio with non-granular cytoplasmic staining
            around the nuclei facilitating easy evaluation of nuclear details [<xref ref-type="bibr" rid="B35">35</xref>]. All 7 SLNs from 7 cases with benign capsular melanocytic nevi showed negative
            results with the intraoperative evaluation of imprint smears.</p>
         <fig id="F2" position="float">
            <label>Figure 2</label>
            <caption>
               <p>Cytomorphological spectrum of tumor cells (arrows) of melanoma metastases immunostained with 'MCW melanoma cocktail'. The tumor cells were larger and showed high N/C ratio with
                  non-granular cytoplasmic staining around the nuclei with usually clear nuclear details. The nuclear chromatin did not simulate the chromatin of lymphocytes in the background. The
                  nucleoli may be prominent. The cell margins were well defined. (HE-FPTS, Hematoxylin-eosin stained formalin-fixed paraffin-embedded tissue sections; ICIS, immunocytochemical
                  evaluation of imprint smears).</p>
            </caption>
            <graphic xlink:href="1746-1596-1-32-2"/>
         </fig>
         <p>An immunostained imprint smear was negative in one case with a small focus of melanoma metastases (4 small groups of 4&#x02013;12 cells each in one small area) in one slice of a single
            SLN. 2 SLNs showed unequivocal melanoma metastases in rapid immunostained imprint smear (Table <xref ref-type="table" rid="T2">2</xref>, case 3 &#x00026; 12). In these 2 SLNs,
            metastases were not detected by immunostained FPTS.</p>
         <p>Two SLNs from 2 patients (Table <xref ref-type="table" rid="T2">2</xref>, case 13 &#x00026; 17) were interpreted as indeterminate. Both of these SLNs were negative for melanoma
            metastases by immunohistochemical evaluation of FPTS. In 2 other cases, rare, singly scattered cells with benign morphology but with cytoplasmic immunostaining with the cocktail were
            observed in both immunostained imprint smears and permanent sections.</p>
      </sec>
      <sec id="id10">
         <title>Discussion</title>
         <p>In the current study, 8 out of 41 patients showed melanoma metastases in lymph nodes employing immunohistochemical evaluation of FPTS. 7 out of these 8 cases showed metastases on imprint
            smears immunostained with the 'MCW melanoma cocktail', potentially avoiding a second surgery at a later date in 87.5% of these patients.</p>
         <p>On a case basis, rapid intraoperative evaluation of imprint smears with the 'MCW melanoma cocktail' yielded a sensitivity of 89% and a specificity of 100%. On a lymph node basis,
            sensitivity was 90% and specificity was 100% (Table <xref ref-type="table" rid="T2">2</xref>). These encouraging results are more promising than alternative intraoperative approaches such
            as the frozen-section alone [<xref ref-type="bibr" rid="B27">27</xref>], immunostaining frozen sections with a cocktail of Melan- A, HMB-45, &#x00026; tyrosinase [<xref ref-type="bibr"
               rid="B32">32</xref>], or a morphological evaluation of imprint smears alone [<xref ref-type="bibr" rid="B30">30</xref>,<xref ref-type="bibr" rid="B38">38</xref>] (Table <xref
               ref-type="table" rid="T3">3</xref>).</p>
         <table-wrap id="T3" position="float">
            <label>Table 3</label>
            <caption>
               <p>Comparison of studies evaluating melanoma SLNs intraoperatively</p>
            </caption>
            <table frame="hsides" rules="groups">
               <thead>
                  <tr>
                     <td align="left">Authors</td>
                     <td align="left">Intraoperative evaluation</td>
                     <td align="center">Sensitivity (%)</td>
                     <td align="center">Specificity (%)</td>
                  </tr>
               </thead>
               <tbody>
                  <tr>
                     <td align="left">Gibbs et al [20A]</td>
                     <td align="left">FS</td>
                     <td align="center">43</td>
                     <td align="center">100</td>
                  </tr>
                  <tr>
                     <td align="left">Clary et al [20B]</td>
                     <td align="left">FS</td>
                     <td align="center">56</td>
                     <td align="center">100</td>
                  </tr>
                  <tr>
                     <td align="left">Tanis et al [20C]</td>
                     <td align="left">FS</td>
                     <td align="center">47</td>
                     <td align="center">100</td>
                  </tr>
                  <tr>
                     <td align="left">Stojadinovic [20]</td>
                     <td align="left">FS</td>
                     <td align="center">59</td>
                     <td align="center">100</td>
                  </tr>
                  <tr>
                     <td align="left">Creager et al [21]</td>
                     <td align="left">IS</td>
                     <td align="center">38</td>
                     <td align="center">100</td>
                  </tr>
                  <tr>
                     <td align="left">Eudy et al [23]</td>
                     <td align="left">FS with IHC</td>
                     <td align="center">86</td>
                     <td align="center">97</td>
                  </tr>
                  <tr>
                     <td align="left">Current study</td>
                     <td align="left">ICIS</td>
                     <td align="center">90</td>
                     <td align="center">100</td>
                  </tr>
               </tbody>
            </table>
            <table-wrap-foot>
               <p>FPTS, formalin-fixed paraffin-embedded tissue section; FS, frozen section, ICIS, immunocytochemical evaluation of imprint smears with 'MCW melanoma cocktail' (mixture of Melan- A,
                  MART- 1, &#x00026; tyrosinase); , IHC, immunohistochemistry with cocktail of Melan- A, HMB-45, &#x00026; tyrosinase); IS, imprint smear; SLNs, sentinel lymph nodes; MM,
                  melanoma metastases.</p>
            </table-wrap-foot>
         </table-wrap>
         <p>Finding a positive SLN and completing the regional lymphadenectomy under the same anesthesia offers many advantages. The extra cost for the second procedure was predominantly the hospital
            cost. In this series with 41 patients, the extra cost (after deducting the cost for the regional lymph node dissection procedure and adding the cost for extended anesthesia) for 8 regional
            lymph node dissections that needed to be performed at a later date was $ 60,000 (average $ 7,500 per case). In comparison, the cost of the intraoperative immunocytochemical evaluation for
            70 SLNs in all 41 cases ([CPT codes 88329 (for intraoperative consult) + 88161 (for preparation and processing of imprint smears for immunostaining) + 88342 (immunostaining and
            interpretation of immunostained smears)] X 70) was only $33,233, resulting in nearly 50% cost saving.</p>
         <p>The most important benefit that cannot be quantified easily is the total avoidance of potential morbidity and mortality associated with an additional surgery at a later date, improving
            patient care. Additionally, saving the patient from a second visit to the surgical suite and its substantial attendant costs along with a decreased number of office visits (to reschedule
            the regional lymphadenectomy), lessens the inconvenience for the patient.</p>
         <p>In cases with positive intraoperative SLN results, the duration of the surgical procedure would be lengthened for the completion of the regional lymphadenectomy. Though it demands
            flexibility in surgical scheduling, it may be worth at most institutions.</p>
         <p>As observed in one case, immunostained imprint smears of SLN that turn out negative may be followed by positive results after immunostaining the FPTS. This may require the patient to
            return to the surgical suite for a lymphadenectomy at a later date. The patients should be made aware of this possibility during preoperative consultations.</p>
         <p>Due to sampling benefits associated with imprint smears [<xref ref-type="bibr" rid="B35">35</xref>], the immunostaining of imprint smears identified metastases that may have been missed by
            studying immunostained FPTS alone. This was observed in 2 cases in which immunostained imprint smears of 2 SLNs were unequivocally positive for melanoma metastases, but immunostained FPTS
            were negative (Table <xref ref-type="table" rid="T2">2</xref>, case 3 &#x00026; 12). Additional SLNs submitted in these 2 patients were positive with both immunostained imprint smears
            and FPTS, further endorsing the true positive nature of finding.</p>
         <p>One SLN from another case showed a micro-focus of melanoma metastases in a single slice in immunostained FPTS. However, the cells from this micro-focus with very few tumor cells were not
            sampled by the imprint smears leading to a false negative result. This underscores the significance of chance factor associated with sampling in general during the evaluation of SLN by any
            method. Unequivocal positive result, obtained with either immunostained imprint smears or with immunostained FPTS, are significant. In cases where the intraoperative results are positive,
            SLNs need not be evaluated further by immunohistochemistry on FPTS. However, if the intraoperative result is negative, immunohistochemical evaluation of SLNs should go forward on FPTS.
            This would enhance the sampling and increase the chances of detecting additional melanoma metastases [<xref ref-type="bibr" rid="B35">35</xref>].</p>
         <p>As a significant advantage, the cells of capsular nevi in 7 SLNs from 7 cases located in lymph node capsule and/or septa as collections of spindle cells, did not exfoliate and adhere to
            slides during the preparation of imprint smears. As discussed previously, this is related to the cohesive nature of benign nevus cells in capsular melanocytic nevi, which would not
            exfoliate cells to the glass slide in contrast to poorly cohesive melanoma cells [<xref ref-type="bibr" rid="B33">33</xref>,<xref ref-type="bibr" rid="B35">35</xref>]. This result in a
            distinct advantage for the immunocytochemical evaluation of imprint smears over other methods such as RT-PCR, which may not have the benefit of a morphological correlation to avoid false
            positive results with benign melanocytic capsular nevi (24).</p>
         <p>Two SLNs from 2 patients, negative for melanoma metastases by immunohistochemical evaluation of FPTS, were interpreted as indeterminate in immunostained imprint smears (Table <xref
               ref-type="table" rid="T2">2</xref> case 13 &#x00026; 17). Retrospectively, the rare doubtful cells observed in immunostained imprint smears were consistent with mast cells which
            demonstrated brown staining (even in respective negative controls) due to endogenous peroxidase activity which could not be blocked during the brief endogenous peroxidase blocking step
            required by the rapid protocol [<xref ref-type="bibr" rid="B35">35</xref>]. Familiarity with morphological spectrum of immunostained tumor cells (Figure <xref ref-type="fig" rid="F1"
            >1</xref>) and other non-specifically stained structures including mast cells in imprint smears prevents indeterminate interpretation [<xref ref-type="bibr" rid="B35">35</xref>]. In two
            other cases, singly scattered cells showing cytoplasmic immunostaining accompanied by small and inconspicuous nuclei were observed. These rare single cells were also present in
            immunostained permanent sections. The cytomorphology was consistent with a benign interpretation in both immunostained imprint smears and the permanent sections. Similar rare cells with
            benign morphology have been reported in SLNs evaluated with MART-1 and Melan-A [<xref ref-type="bibr" rid="B39">39</xref>].</p>
         <p>For various reasons every laboratory may not be comfortable using cytological methods. However, objective nature of interpreting immunostained tumor cells in imprint smears would allow
            interpretation by most of the pathology laboratories after a minor learning curve [<xref ref-type="bibr" rid="B35">35</xref>].</p>
         <p>Imprint smears are less expensive and faster than frozen sectioning without cryostat related tissue loss. They avoid the frequently observed problems associated with the frozen sectioning
            of fatty lymph nodes. If imprint smears are made and the tissue is not frozen, it would prevent the introduction of freezing artifacts and the associated interpretation challenges on the
            permanent sections of such frozen tissue.</p>
         <p>Other approaches for the intraoperative evaluation of SLN have demonstrated lower sensitivities (Table <xref ref-type="table" rid="T3">3</xref>) but had 100% specificity, which is
            extremely important in order to avoid un-indicated regional lymph-adenectomies [<xref ref-type="bibr" rid="B26">26</xref>-<xref ref-type="bibr" rid="B32">32</xref>]. One study evaluating
            immunostaining of frozen sections demonstrated a slightly increased sensitivity than when imprint smears or frozen sections alone were used without immunostaining. However, the specificity
            was less than 100% (Table <xref ref-type="table" rid="T3">3</xref>). The immunostained frozen sections may show significant artifacts along with folds, background staining, missing areas,
            and poor morphology. As HMB45 is known to stain non-melanoma cells in SLN [<xref ref-type="bibr" rid="B33">33</xref>,<xref ref-type="bibr" rid="B40">40</xref>-<xref ref-type="bibr"
               rid="B42">42</xref>], incorporation of HMB45 in the cocktail used in this study may have contributed further to a reduction in the specificity [<xref ref-type="bibr" rid="B32"
               >32</xref>,<xref ref-type="bibr" rid="B41">41</xref>,<xref ref-type="bibr" rid="B42">42</xref>]. As stated by the reporting authors of the study, a recent consensus group has
            discouraged frozen-section examination of SLN [<xref ref-type="bibr" rid="B32">32</xref>].</p>
         <p>In summary, imprint smears, rapidly immunostained with the 'MCW melanoma cocktail' is a sensitive and specific method for the rapid intraoperative evaluation of SLNs. If a SLN is found to
            be positive for melanoma metastases in this way, the completion of a regional lymphadenectomy during the same surgical procedure for SLN biopsy is feasible. Both negative and indeterminate
            results should be regarded as negative when considering intraoperative decision making for a regional lymph node dissection during a given surgical procedure. In such cases, the final
            decision for a regional lymph node dissection should be deferred until the final results with FPTS are available.</p>
      </sec>
      <sec id="id11">
         <title>Abbreviations</title>
         <p>FPTS, formalin-fixed paraffin-embedded tissue section; H&#x00026;E, hematoxylin and eosin; SLN, sentinel lymph node;</p>
      </sec>
   </body>
   <back>
      <ack>
         <sec id="id12">
            <title>Acknowledgements</title>
            <p>Part of this study was supported by the Cancer Center Research Multidisciplinary Grant from Medical College of Wisconsin, Milwaukee, WI and in part by Armour family grant through
               Chicago Marathon.</p>
            <p>Authors thank Virgilia Macias, MD; Sushma Kaul, MS; Glen Dawson, BS,HT,QIHC(ASCP); Aurora Kallenbach, HT(ASCP); and Jerome Jacobson, HT, QIHC(ASCP) for their expert technical assistance
               in immunochemistry. We also thank Christine M. Santiago, Compliance Education Coordinator at MCW for reviewing the cost and CPT code related comments.</p>
         </sec>
      </ack>
      <ref-list>
         <ref id="B1">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Landis</surname>
                     <given-names>SH</given-names>
                  </name>
                  <name>
                     <surname>Murray</surname>
                     <given-names>T</given-names>
                  </name>
                  <name>
                     <surname>Bolden</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Wingo</surname>
                     <given-names>PA</given-names>
                  </name>
               </person-group>
               <article-title>Cancer statistics, 1999</article-title>
               <source>CA Cancer J Clin</source>
               <year>1999</year>
               <volume>49</volume>
               <fpage>8</fpage>
               <lpage>31,1</lpage>
               <pub-id pub-id-type="pmid">10200775</pub-id>
            </citation>
         </ref>
         <ref id="B2">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Medalie</surname>
                     <given-names>NS</given-names>
                  </name>
                  <name>
                     <surname>Ackerman</surname>
                     <given-names>AB</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel lymph node biopsy has no benefit for patients with primary cutaneous melanoma metastatic to a lymph node: an assertion based on comprehensive, critical analysis:
                  part I</article-title>
               <source>Am J Dermatopathol</source>
               <year>2003</year>
               <volume>25</volume>
               <fpage>399</fpage>
               <lpage>417</lpage>
               <pub-id pub-id-type="pmid">14501289</pub-id>
            </citation>
         </ref>
         <ref id="B3">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Medalie</surname>
                     <given-names>NS</given-names>
                  </name>
                  <name>
                     <surname>Ackerman</surname>
                     <given-names>AB</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel lymph node biopsy has no benefit for patients with primary cutaneous melanoma metastatic to a lymph node: an assertion based on comprehensive, critical analysis:
                  part II</article-title>
               <source>Am J Dermatopathol</source>
               <year>2003</year>
               <volume>25</volume>
               <fpage>473</fpage>
               <lpage>84</lpage>
               <pub-id pub-id-type="pmid">14631188</pub-id>
            </citation>
         </ref>
         <ref id="B4">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Balch</surname>
                     <given-names>CM</given-names>
                  </name>
                  <name>
                     <surname>Soong</surname>
                     <given-names>SJ</given-names>
                  </name>
                  <name>
                     <surname>Bartolucci</surname>
                     <given-names>AA</given-names>
                  </name>
                  <name>
                     <surname>Urist</surname>
                     <given-names>MM</given-names>
                  </name>
                  <name>
                     <surname>Karakousis</surname>
                     <given-names>CP</given-names>
                  </name>
                  <name>
                     <surname>Smith</surname>
                     <given-names>TJ</given-names>
                  </name>
                  <name>
                     <surname>Temple</surname>
                     <given-names>WJ</given-names>
                  </name>
                  <name>
                     <surname>Ross</surname>
                     <given-names>MI</given-names>
                  </name>
                  <name>
                     <surname>Jewell</surname>
                     <given-names>WR</given-names>
                  </name>
                  <name>
                     <surname>Mihm</surname>
                     <given-names>MC</given-names>
                  </name>
                  <name>
                     <surname>Barnhill</surname>
                     <given-names>RL</given-names>
                  </name>
                  <name>
                     <surname>Wanebo</surname>
                     <given-names>HJ</given-names>
                  </name>
               </person-group>
               <article-title>Efficacy of an elective regional lymph node dissection of 1 to 4 mm thick melanomas for patients 60 years of age and younger</article-title>
               <source>Ann Surg</source>
               <year>1996</year>
               <volume>224</volume>
               <fpage>255</fpage>
               <lpage>63</lpage>
               <comment>discussion 263&#x02013;6.</comment>
               <pub-id pub-id-type="pmid">8813254</pub-id>
               <pub-id pub-id-type="doi">10.1097/00000658-199609000-00002</pub-id>
            </citation>
         </ref>
         <ref id="B5">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Cascinelli</surname>
                     <given-names>N</given-names>
                  </name>
                  <name>
                     <surname>Morabito</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Santinami</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>MacKie</surname>
                     <given-names>RM</given-names>
                  </name>
                  <name>
                     <surname>Belli</surname>
                     <given-names>F</given-names>
                  </name>
               </person-group>
               <article-title>Immediate or delayed dissection of regional nodes in patients with melanoma of the trunk: a randomised trial</article-title>
               <source>WHO Melanoma Programme Lancet</source>
               <year>1998</year>
               <volume>351</volume>
               <fpage>793</fpage>
               <lpage>6</lpage>
            </citation>
         </ref>
         <ref id="B6">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Salti</surname>
                     <given-names>GI</given-names>
                  </name>
                  <name>
                     <surname>Das Gupta</surname>
                     <given-names>TK</given-names>
                  </name>
               </person-group>
               <article-title>Predicting residual lymph node basin disease in melanoma patients with sentinel lymph node metastases</article-title>
               <source>Am J Surg</source>
               <year>2003</year>
               <volume>186</volume>
               <fpage>98</fpage>
               <lpage>101</lpage>
               <pub-id pub-id-type="pmid">12885597</pub-id>
               <pub-id pub-id-type="doi">10.1016/S0002-9610(03)00169-7</pub-id>
            </citation>
         </ref>
         <ref id="B7">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Nowecki</surname>
                     <given-names>ZI</given-names>
                  </name>
                  <name>
                     <surname>Rutkowski</surname>
                     <given-names>P</given-names>
                  </name>
                  <name>
                     <surname>Nasierowska-Guttmejer</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Ruka</surname>
                     <given-names>W</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel lymph node biopsy in melanoma patients with clinically negative regional lymph nodes &#x02013; one institution's experience</article-title>
               <source>Melanoma Res</source>
               <year>2003</year>
               <volume>13</volume>
               <fpage>35</fpage>
               <lpage>43</lpage>
               <pub-id pub-id-type="pmid">12569283</pub-id>
               <pub-id pub-id-type="doi">10.1097/00008390-200302000-00007</pub-id>
            </citation>
         </ref>
         <ref id="B8">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Manca</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Facchetti</surname>
                     <given-names>F</given-names>
                  </name>
                  <name>
                     <surname>Pizzocaro</surname>
                     <given-names>C</given-names>
                  </name>
                  <name>
                     <surname>Biasca</surname>
                     <given-names>F</given-names>
                  </name>
                  <name>
                     <surname>Farfaglia</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>Simoncini</surname>
                     <given-names>E</given-names>
                  </name>
                  <name>
                     <surname>Cristinelli</surname>
                     <given-names>MR</given-names>
                  </name>
                  <name>
                     <surname>Flocchini</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Parrinello</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Manganoni</surname>
                     <given-names>A</given-names>
                  </name>
               </person-group>
               <article-title>Nodal staging in localized melanoma. The experience of the Brescia Melanoma Unit</article-title>
               <source>Br J Plast Surg</source>
               <year>2003</year>
               <volume>56</volume>
               <fpage>534</fpage>
               <lpage>9</lpage>
               <pub-id pub-id-type="pmid">12946370</pub-id>
               <pub-id pub-id-type="doi">10.1016/S0007-1226(03)00208-X</pub-id>
            </citation>
         </ref>
         <ref id="B9">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Rutkowski</surname>
                     <given-names>P</given-names>
                  </name>
                  <name>
                     <surname>Nowecki</surname>
                     <given-names>ZI</given-names>
                  </name>
                  <name>
                     <surname>Nasierowska-Guttmejer</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Ruka</surname>
                     <given-names>W</given-names>
                  </name>
               </person-group>
               <article-title>Lymph node status and survival in cutaneous malignant melanoma &#x02013; sentinel lymph node biopsy impact</article-title>
               <source>Eur J Surg Oncol</source>
               <year>2003</year>
               <volume>29</volume>
               <fpage>611</fpage>
               <lpage>8</lpage>
               <pub-id pub-id-type="pmid">12943629</pub-id>
               <pub-id pub-id-type="doi">10.1016/S0748-7983(03)00118-5</pub-id>
            </citation>
         </ref>
         <ref id="B10">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Wagner</surname>
                     <given-names>JD</given-names>
                  </name>
                  <name>
                     <surname>Ranieri</surname>
                     <given-names>J</given-names>
                  </name>
                  <name>
                     <surname>Evdokimow</surname>
                     <given-names>DZ</given-names>
                  </name>
                  <name>
                     <surname>Logan</surname>
                     <given-names>T</given-names>
                  </name>
                  <name>
                     <surname>Chuang</surname>
                     <given-names>TY</given-names>
                  </name>
                  <name>
                     <surname>Johnson</surname>
                     <given-names>CS</given-names>
                  </name>
                  <name>
                     <surname>Jung</surname>
                     <given-names>SH</given-names>
                  </name>
                  <name>
                     <surname>Wenck</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Coleman</surname>
                     <given-names>JJ</given-names>
                     <suffix>3rd</suffix>
                  </name>
               </person-group>
               <article-title>Patterns of initial recurrence and prognosis after sentinel lymph node biopsy and selective lymphadenectomy for melanoma</article-title>
               <source>Plast Reconstr Surg</source>
               <year>2003</year>
               <volume>112</volume>
               <fpage>486</fpage>
               <lpage>97</lpage>
               <pub-id pub-id-type="pmid">12900606</pub-id>
               <pub-id pub-id-type="doi">10.1097/01.PRS.0000070989.23469.1F</pub-id>
            </citation>
         </ref>
         <ref id="B11">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Zapas</surname>
                     <given-names>JL</given-names>
                  </name>
                  <name>
                     <surname>Coley</surname>
                     <given-names>HC</given-names>
                  </name>
                  <name>
                     <surname>Beam</surname>
                     <given-names>SL</given-names>
                  </name>
                  <name>
                     <surname>Brown</surname>
                     <given-names>SD</given-names>
                  </name>
                  <name>
                     <surname>Jablonski</surname>
                     <given-names>KA</given-names>
                  </name>
                  <name>
                     <surname>Elias</surname>
                     <given-names>EG</given-names>
                  </name>
               </person-group>
               <article-title>The risk of regional lymph node metastases in patients with melanoma less than 1.0 mm thick: recommendations for sentinel lymph node biopsy</article-title>
               <source>J Am Coll Surg</source>
               <year>2003</year>
               <volume>197</volume>
               <fpage>403</fpage>
               <lpage>7</lpage>
               <pub-id pub-id-type="pmid">12946795</pub-id>
               <pub-id pub-id-type="doi">10.1016/S1072-7515(03)00432-0</pub-id>
            </citation>
         </ref>
         <ref id="B12">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Gad</surname>
                     <given-names>D</given-names>
                  </name>
                  <name>
                     <surname>Sjogren</surname>
                     <given-names>P</given-names>
                  </name>
               </person-group>
               <article-title>[Sentinel lymph node biopsy in patients with cutaneous malignant melanoma]</article-title>
               <source>Ugeskr Laeger</source>
               <year>2003</year>
               <volume>165</volume>
               <fpage>2567</fpage>
               <lpage>70</lpage>
               <comment>Danish.</comment>
               <pub-id pub-id-type="pmid">12854268</pub-id>
            </citation>
         </ref>
         <ref id="B13">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Kaikaris</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Valiukeviciene</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Rimdeika</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>Gollnick</surname>
                     <given-names>H</given-names>
                  </name>
                  <name>
                     <surname>Ulrich</surname>
                     <given-names>J</given-names>
                  </name>
               </person-group>
               <article-title>[Sentinel lymph node biopsy in melanoma patients: methods, indications, and clinical significance]</article-title>
               <source>Medicina (Kaunas)</source>
               <year>2003</year>
               <volume>39</volume>
               <fpage>621</fpage>
               <lpage>30</lpage>
               <comment>Review. Lithuanian.</comment>
               <pub-id pub-id-type="pmid">12878814</pub-id>
            </citation>
         </ref>
         <ref id="B14">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Arens</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Osinga</surname>
                     <given-names>J</given-names>
                  </name>
                  <name>
                     <surname>Schwipper</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Schober</surname>
                     <given-names>O</given-names>
                  </name>
                  <name>
                     <surname>Tilkorn</surname>
                     <given-names>H</given-names>
                  </name>
                  <name>
                     <surname>Liebau</surname>
                     <given-names>J</given-names>
                  </name>
               </person-group>
               <article-title>[Sentinel lymph node dissection in patients with malignant melanoma. Diagnostic and therapeutic standards]</article-title>
               <source>Chirurg</source>
               <year>2003</year>
               <volume>74</volume>
               <fpage>665</fpage>
               <lpage>70</lpage>
               <comment>German.</comment>
               <pub-id pub-id-type="pmid">12883795</pub-id>
               <pub-id pub-id-type="doi">10.1007/s00104-003-0659-x</pub-id>
            </citation>
         </ref>
         <ref id="B15">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Santinami</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Baldi</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Tragni</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Maurichi</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Moglia</surname>
                     <given-names>D</given-names>
                  </name>
                  <name>
                     <surname>Bono</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Bartoli</surname>
                     <given-names>C</given-names>
                  </name>
                  <name>
                     <surname>Patuzzo</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>Pennacchioli</surname>
                     <given-names>E</given-names>
                  </name>
                  <name>
                     <surname>Cascinelli</surname>
                     <given-names>N</given-names>
                  </name>
               </person-group>
               <article-title>[Impact of sentinel lymph node biopsy on the treatment of melanoma]</article-title>
               <source>Tumori</source>
               <year>2003</year>
               <volume>89</volume>
               <fpage>273</fpage>
               <lpage>5</lpage>
               <comment>Italian.</comment>
               <pub-id pub-id-type="pmid">12903619</pub-id>
            </citation>
         </ref>
         <ref id="B16">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Al-Hiari</surname>
                     <given-names>AA</given-names>
                  </name>
                  <name>
                     <surname>Al-Kaylani</surname>
                     <given-names>H</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel node in management of malignant melanoma</article-title>
               <source>Saudi Med J</source>
               <year>2002</year>
               <volume>23</volume>
               <fpage>1462</fpage>
               <lpage>5</lpage>
               <pub-id pub-id-type="pmid">12518192</pub-id>
            </citation>
         </ref>
         <ref id="B17">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Pu</surname>
                     <given-names>LL</given-names>
                  </name>
                  <name>
                     <surname>Wells</surname>
                     <given-names>KE</given-names>
                  </name>
                  <name>
                     <surname>Cruse</surname>
                     <given-names>CW</given-names>
                  </name>
                  <name>
                     <surname>Shons</surname>
                     <given-names>AR</given-names>
                  </name>
                  <name>
                     <surname>Reintgen</surname>
                     <given-names>DS</given-names>
                  </name>
               </person-group>
               <article-title>Prevalence of additional positive lymph nodes in complete lymphadenectomy specimens after positive sentinel lymphadenectomy findings for early-stage melanoma of the head
                  and neck</article-title>
               <source>Plast Reconstr Surg</source>
               <year>2003</year>
               <volume>112</volume>
               <fpage>43</fpage>
               <lpage>9</lpage>
               <pub-id pub-id-type="pmid">12832875</pub-id>
               <pub-id pub-id-type="doi">10.1097/01.PRS.0000065912.20180.A9</pub-id>
            </citation>
         </ref>
         <ref id="B18">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Buonomo</surname>
                     <given-names>O</given-names>
                  </name>
                  <name>
                     <surname>Felici</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Granai</surname>
                     <given-names>AV</given-names>
                  </name>
                  <name>
                     <surname>Piccirillo</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>De Liguori Carino</surname>
                     <given-names>N</given-names>
                  </name>
                  <name>
                     <surname>Guadagni</surname>
                     <given-names>F</given-names>
                  </name>
                  <name>
                     <surname>Mariotti</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Orlandi</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Tipaldi</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Cipriani</surname>
                     <given-names>C</given-names>
                  </name>
                  <name>
                     <surname>Chimenti</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Cervelli</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Casciani</surname>
                     <given-names>CU</given-names>
                  </name>
                  <name>
                     <surname>Roselli</surname>
                     <given-names>M</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel lymphadenectomy in cutaneous melanoma</article-title>
               <source>Tumori</source>
               <year>2002</year>
               <volume>88</volume>
               <fpage>S49</fpage>
               <lpage>51</lpage>
               <pub-id pub-id-type="pmid">12369552</pub-id>
            </citation>
         </ref>
         <ref id="B19">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Nieweg</surname>
                     <given-names>OE</given-names>
                  </name>
                  <name>
                     <surname>Eggermont</surname>
                     <given-names>AM</given-names>
                  </name>
                  <name>
                     <surname>Kroon</surname>
                     <given-names>BB</given-names>
                  </name>
               </person-group>
               <article-title>[Sentinel node biopsy in melanoma: some critical questions]</article-title>
               <source>Ned Tijdschr Geneeskd</source>
               <year>2002</year>
               <volume>146</volume>
               <fpage>1774</fpage>
               <lpage>7</lpage>
               <comment>Review. Dutch</comment>
               <pub-id pub-id-type="pmid">12369437</pub-id>
            </citation>
         </ref>
         <ref id="B20">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Essner</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>Cochran</surname>
                     <given-names>AJ</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel node biopsy: not only a staging tool?</article-title>
               <source>Recent Results Cancer Res</source>
               <year>2002</year>
               <volume>160</volume>
               <fpage>133</fpage>
               <lpage>48</lpage>
               <pub-id pub-id-type="pmid">12079207</pub-id>
            </citation>
         </ref>
         <ref id="B21">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Thompson</surname>
                     <given-names>JF</given-names>
                  </name>
               </person-group>
               <article-title>The Sydney Melanoma Unit experience of sentinel lymphadenectomy for melanoma</article-title>
               <source>Ann Surg Oncol</source>
               <year>2001</year>
               <volume>8</volume>
               <fpage>44S</fpage>
               <lpage>47S</lpage>
               <pub-id pub-id-type="pmid">11599898</pub-id>
            </citation>
         </ref>
         <ref id="B22">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Cascinelli</surname>
                     <given-names>N</given-names>
                  </name>
                  <name>
                     <surname>Belli</surname>
                     <given-names>F</given-names>
                  </name>
                  <name>
                     <surname>Santinami</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Fait</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Testori</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Ruka</surname>
                     <given-names>W</given-names>
                  </name>
                  <name>
                     <surname>Cavaliere</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>Mozzillo</surname>
                     <given-names>N</given-names>
                  </name>
                  <name>
                     <surname>Rossi</surname>
                     <given-names>CR</given-names>
                  </name>
                  <name>
                     <surname>MacKie</surname>
                     <given-names>RM</given-names>
                  </name>
                  <name>
                     <surname>Nieweg</surname>
                     <given-names>O</given-names>
                  </name>
                  <name>
                     <surname>Pace</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Kirov</surname>
                     <given-names>K</given-names>
                  </name>
               </person-group>
               <article-title>Sentinel lymph node biopsy in cutaneous melanoma: the WHO Melanoma Program experience</article-title>
               <source>Ann Surg Oncol</source>
               <year>2000</year>
               <volume>7</volume>
               <fpage>469</fpage>
               <lpage>74</lpage>
               <pub-id pub-id-type="pmid">10894144</pub-id>
               <pub-id pub-id-type="doi">10.1007/s10434-000-0469-z</pub-id>
            </citation>
         </ref>
         <ref id="B23">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Havenga</surname>
                     <given-names>K</given-names>
                  </name>
                  <name>
                     <surname>Cobben</surname>
                     <given-names>DC</given-names>
                  </name>
                  <name>
                     <surname>Oyen</surname>
                     <given-names>WJ</given-names>
                  </name>
                  <name>
                     <surname>Nienhuijs</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Hoekstra</surname>
                     <given-names>HJ</given-names>
                  </name>
                  <name>
                     <surname>Ruers</surname>
                     <given-names>TJ</given-names>
                  </name>
                  <name>
                     <surname>Wobbes</surname>
                     <given-names>T</given-names>
                  </name>
               </person-group>
               <article-title>Fluorodeoxyglucose-positron emission tomography and sentinel lymph node biopsy in staging primary cutaneous melanoma</article-title>
               <source>Eur J Surg Oncol</source>
               <year>2003</year>
               <volume>29</volume>
               <fpage>662</fpage>
               <lpage>4</lpage>
               <pub-id pub-id-type="pmid">14511614</pub-id>
               <pub-id pub-id-type="doi">10.1016/S0748-7983(03)00147-1</pub-id>
            </citation>
         </ref>
         <ref id="B24">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Schafer</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Herbst</surname>
                     <given-names>RA</given-names>
                  </name>
                  <name>
                     <surname>Beiteke</surname>
                     <given-names>U</given-names>
                  </name>
                  <name>
                     <surname>Lange-Ionescu</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Treckmann</surname>
                     <given-names>H</given-names>
                  </name>
                  <name>
                     <surname>Lohlein</surname>
                     <given-names>D</given-names>
                  </name>
                  <name>
                     <surname>Thiemann</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Theophil</surname>
                     <given-names>B</given-names>
                  </name>
                  <name>
                     <surname>Schwarze</surname>
                     <given-names>EW</given-names>
                  </name>
                  <name>
                     <surname>Bartels</surname>
                     <given-names>HJ</given-names>
                  </name>
                  <name>
                     <surname>Frosch</surname>
                     <given-names>PJ</given-names>
                  </name>
               </person-group>
               <article-title>[Sentinel lymph node excision (SLNE) and positron emission tomography in the staging of stage I-II melanoma patients]</article-title>
               <source>Hautarzt</source>
               <year>2003</year>
               <volume>54</volume>
               <fpage>440</fpage>
               <lpage>7</lpage>
               <comment>German.</comment>
               <pub-id pub-id-type="pmid">12719864</pub-id>
            </citation>
         </ref>
         <ref id="B25">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Stojadinovic</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Allen</surname>
                     <given-names>PJ</given-names>
                  </name>
                  <name>
                     <surname>Clary</surname>
                     <given-names>BM</given-names>
                  </name>
                  <name>
                     <surname>Busam</surname>
                     <given-names>KJ</given-names>
                  </name>
                  <name>
                     <surname>Coit</surname>
                     <given-names>DG</given-names>
                  </name>
               </person-group>
               <article-title>Value of frozen-section analysis of sentinel lymph nodes for primary cutaneous malignant melanoma</article-title>
               <source>Ann Surg</source>
               <year>2002</year>
               <volume>235</volume>
               <fpage>92</fpage>
               <lpage>8</lpage>
               <pub-id pub-id-type="pmid">11753047</pub-id>
               <pub-id pub-id-type="doi">10.1097/00000658-200201000-00012</pub-id>
            </citation>
         </ref>
         <ref id="B26">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Gibbs</surname>
                     <given-names>JF</given-names>
                  </name>
                  <name>
                     <surname>Huang</surname>
                     <given-names>PP</given-names>
                  </name>
                  <name>
                     <surname>Zhang</surname>
                     <given-names>PJ</given-names>
                  </name>
                  <name>
                     <surname>Kraybill</surname>
                     <given-names>WG</given-names>
                  </name>
                  <name>
                     <surname>Cheney</surname>
                     <given-names>R</given-names>
                  </name>
               </person-group>
               <article-title>Accuracy of pathologic techniques for the diagnosis of metastatic melanoma in sentinel lymph nodes</article-title>
               <source>Ann Surg Oncol</source>
               <year>1999</year>
               <volume>6</volume>
               <fpage>699</fpage>
               <lpage>704</lpage>
               <pub-id pub-id-type="pmid">10560857</pub-id>
            </citation>
         </ref>
         <ref id="B27">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Clary</surname>
                     <given-names>BM</given-names>
                  </name>
                  <name>
                     <surname>Lewis</surname>
                     <given-names>JJ</given-names>
                  </name>
                  <name>
                     <surname>Brady</surname>
                     <given-names>MS</given-names>
                  </name>
                  <name>
                     <surname>Busam</surname>
                     <given-names>K</given-names>
                  </name>
                  <name>
                     <surname>Coit</surname>
                     <given-names>DG</given-names>
                  </name>
               </person-group>
               <article-title>Should frozen section analysis of the sentinel node be performed in patients with melanoma?</article-title>
               <source>Eur J Nucl Med</source>
               <year>1999</year>
               <volume>26</volume>
               <fpage>S68</fpage>
            </citation>
         </ref>
         <ref id="B28">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Tanis</surname>
                     <given-names>PJ</given-names>
                  </name>
                  <name>
                     <surname>Boom</surname>
                     <given-names>RP</given-names>
                  </name>
                  <name>
                     <surname>Koops</surname>
                     <given-names>HS</given-names>
                  </name>
                  <name>
                     <surname>Faneyte</surname>
                     <given-names>IF</given-names>
                  </name>
                  <name>
                     <surname>Peterse</surname>
                     <given-names>JL</given-names>
                  </name>
                  <name>
                     <surname>Nieweg</surname>
                     <given-names>OE</given-names>
                  </name>
                  <name>
                     <surname>Rutgers</surname>
                     <given-names>EJ</given-names>
                  </name>
                  <name>
                     <surname>Tiebosch</surname>
                     <given-names>AT</given-names>
                  </name>
                  <name>
                     <surname>Kroon</surname>
                     <given-names>BB</given-names>
                  </name>
               </person-group>
               <article-title>Frozen section investigation of the sentinel node in malignant melanoma and breast cancer</article-title>
               <source>Ann Surg Oncol</source>
               <year>2001</year>
               <volume>8</volume>
               <fpage>222</fpage>
               <lpage>26</lpage>
               <pub-id pub-id-type="pmid">11314938</pub-id>
            </citation>
         </ref>
         <ref id="B29">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Creager</surname>
                     <given-names>AJ</given-names>
                  </name>
                  <name>
                     <surname>Shiver</surname>
                     <given-names>SA</given-names>
                  </name>
                  <name>
                     <surname>Shen</surname>
                     <given-names>P</given-names>
                  </name>
                  <name>
                     <surname>Geisinger</surname>
                     <given-names>KR</given-names>
                  </name>
                  <name>
                     <surname>Levine</surname>
                     <given-names>EA</given-names>
                  </name>
               </person-group>
               <article-title>Intraoperative evaluation of sentinel lymph nodes for metastatic melanoma by imprint cytology</article-title>
               <source>Cancer</source>
               <year>2002</year>
               <volume>94</volume>
               <fpage>3016</fpage>
               <lpage>22</lpage>
               <pub-id pub-id-type="pmid">12115392</pub-id>
               <pub-id pub-id-type="doi">10.1002/cncr.10512</pub-id>
            </citation>
         </ref>
         <ref id="B30">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Hocevat</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Bracko</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Pogacnik</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Vidergar-Kralj</surname>
                     <given-names>B</given-names>
                  </name>
                  <name>
                     <surname>Besic</surname>
                     <given-names>N</given-names>
                  </name>
                  <name>
                     <surname>Zgajnar</surname>
                     <given-names>J</given-names>
                  </name>
               </person-group>
               <article-title>Role of imprint cytology in the intraoperative evaluation of sentinel lymph nodes for malignant melanoma</article-title>
               <source>Eur J Cancer</source>
               <year>2003</year>
               <volume>39</volume>
               <fpage>2173</fpage>
               <lpage>8</lpage>
               <pub-id pub-id-type="pmid">14522375</pub-id>
               <pub-id pub-id-type="doi">10.1016/S0959-8049(03)00453-2</pub-id>
            </citation>
         </ref>
         <ref id="B31">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Eudy</surname>
                     <given-names>GE</given-names>
                  </name>
                  <name>
                     <surname>Carlson</surname>
                     <given-names>GW</given-names>
                  </name>
                  <name>
                     <surname>Murray</surname>
                     <given-names>DR</given-names>
                  </name>
                  <name>
                     <surname>Waldrop</surname>
                     <given-names>SM</given-names>
                  </name>
                  <name>
                     <surname>Lawson</surname>
                     <given-names>D</given-names>
                  </name>
                  <name>
                     <surname>Cohen</surname>
                     <given-names>C</given-names>
                  </name>
               </person-group>
               <article-title>Rapid immunohistochemistry of sentinel lymph nodes for metastatic melanoma</article-title>
               <source>Hum Pathol</source>
               <year>2003</year>
               <volume>34</volume>
               <fpage>797</fpage>
               <lpage>802</lpage>
               <pub-id pub-id-type="pmid">14506642</pub-id>
               <pub-id pub-id-type="doi">10.1016/S0046-8177(03)00290-9</pub-id>
            </citation>
         </ref>
         <ref id="B32">
            <citation citation-type="other">
               <person-group person-group-type="author">
                  <name>
                     <surname>Shidham</surname>
                     <given-names>VB</given-names>
                  </name>
                  <name>
                     <surname>Qi</surname>
                     <given-names>D</given-names>
                  </name>
                  <name>
                     <surname>Rao</surname>
                     <given-names>RN</given-names>
                  </name>
                  <name>
                     <surname>Acker</surname>
                     <given-names>SM</given-names>
                  </name>
                  <name>
                     <surname>Chang</surname>
                     <given-names>CC</given-names>
                  </name>
                  <name>
                     <surname>Kampalath</surname>
                     <given-names>B</given-names>
                  </name>
                  <name>
                     <surname>Dawson</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Machhi</surname>
                     <given-names>JK</given-names>
                  </name>
                  <name>
                     <surname>Komorowski</surname>
                     <given-names>RA</given-names>
                  </name>
               </person-group>
               <article-title>Improved Immunohistochemical Evaluation of Micrometastases in Sentinel Lymph Nodes of Cutaneous Melanoma with MCW Melanoma Cocktail- A Mixture of Monoclonal Antibodies to
                  MART-1, Melan-A, and Tyrosinase</article-title>
               <source>BMC Cancer</source>
               <volume>3</volume>
               <fpage>p1</fpage>
               <lpage>9</lpage>
               <ext-link ext-link-type="uri" xlink:href="http://www.biomedcentral.com/qc/1471-2407/3/15"/>
               <comment>2003 May 7;[PubMed PMID: 12735792]</comment>
            </citation>
         </ref>
         <ref id="B33">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Shidham</surname>
                     <given-names>VB</given-names>
                  </name>
                  <name>
                     <surname>Komorowski</surname>
                     <given-names>R</given-names>
                  </name>
               </person-group>
               <article-title>'MCW melanoma cocktail' for evaluation of metastases in sentinel lymph nodes of cutaneous melanoma</article-title>
               <source>Expert Review of Molecular Diagnostics</source>
               <year>2005</year>
               <volume>5</volume>
               <fpage>281</fpage>
               <lpage>290</lpage>
               <pub-id pub-id-type="pmid">15934808</pub-id>
               <pub-id pub-id-type="doi">10.1586/14737159.5.3.281</pub-id>
            </citation>
         </ref>
         <ref id="B34">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Shidham</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Macias</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Neuberg</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Chavan</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Rao</surname>
                     <given-names>RN</given-names>
                  </name>
                  <name>
                     <surname>Basir</surname>
                     <given-names>Z</given-names>
                  </name>
                  <name>
                     <surname>Walker</surname>
                     <given-names>A</given-names>
                  </name>
                  <name>
                     <surname>Dzwierzynski</surname>
                     <given-names>WW</given-names>
                  </name>
               </person-group>
               <article-title>Rapid intraoperative immunocytochemical evaluation of cutaneous melanoma sentinel lymph nodes for melanoma metastases with 'MCW Melanoma Cocktail'</article-title>
               <source>Acta Cytol</source>
               <year>2003</year>
               <volume>47</volume>
               <fpage>938</fpage>
            </citation>
         </ref>
         <ref id="B35">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Shidham</surname>
                     <given-names>VB</given-names>
                  </name>
                  <name>
                     <surname>Komorowski</surname>
                     <given-names>R</given-names>
                  </name>
                  <name>
                     <surname>Dawson</surname>
                     <given-names>G</given-names>
                  </name>
                  <name>
                     <surname>Kaul</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Chang</surname>
                     <given-names>CC</given-names>
                  </name>
                  <name>
                     <surname>Macias</surname>
                     <given-names>V</given-names>
                  </name>
               </person-group>
               <article-title>Optimization of immunostaining protocol for rapid intraoperative evaluation of imprint smears of melanoma sentinel lymph nodes with 'MCW melanoma cocktail'</article-title>
               <source>CytoJournal</source>
               <year>2004</year>
               <volume>1</volume>
               <fpage>2</fpage>
               <ext-link ext-link-type="uri" xlink:href="http://www.cytojournal.com/content/1/1/2"/>
               <comment>doi:10.1186/1742-6413-1-2</comment>
               <pub-id pub-id-type="pmid">15500702</pub-id>
               <pub-id pub-id-type="doi">10.1186/1742-6413-1-2</pub-id>
            </citation>
         </ref>
         <ref id="B36">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Spanknebel</surname>
                     <given-names>K</given-names>
                  </name>
                  <name>
                     <surname>Coit</surname>
                     <given-names>DG</given-names>
                  </name>
                  <name>
                     <surname>Bieligk</surname>
                     <given-names>SC</given-names>
                  </name>
                  <name>
                     <surname>Gonen</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Rosai</surname>
                     <given-names>J</given-names>
                  </name>
                  <name>
                     <surname>Klimstra</surname>
                     <given-names>DS</given-names>
                  </name>
               </person-group>
               <article-title>Characterization of Micrometastatic Disease in Melanoma Sentinel Lymph Nodes by Enhanced Pathology: Recommendations for Standardizing Pathologic Analysis</article-title>
               <source>Am J Surg Pathol</source>
               <year>2005</year>
               <volume>29</volume>
               <fpage>305</fpage>
               <lpage>17</lpage>
               <pub-id pub-id-type="pmid">15725798</pub-id>
               <pub-id pub-id-type="doi">10.1097/01.pas.0000152134.36030.b7</pub-id>
            </citation>
         </ref>
         <ref id="B37">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Kapteijn</surname>
                     <given-names>BA</given-names>
                  </name>
                  <name>
                     <surname>Nieweg</surname>
                     <given-names>OE</given-names>
                  </name>
                  <name>
                     <surname>Liem</surname>
                     <given-names>I</given-names>
                  </name>
                  <name>
                     <surname>Mooi</surname>
                     <given-names>WJ</given-names>
                  </name>
                  <name>
                     <surname>Balm</surname>
                     <given-names>AJ</given-names>
                  </name>
                  <name>
                     <surname>Muller</surname>
                     <given-names>SH</given-names>
                  </name>
                  <name>
                     <surname>Peterse</surname>
                     <given-names>JL</given-names>
                  </name>
                  <name>
                     <surname>Valdes Olmos</surname>
                     <given-names>RA</given-names>
                  </name>
                  <name>
                     <surname>Hoefnagel</surname>
                     <given-names>CA</given-names>
                  </name>
                  <name>
                     <surname>Kroon</surname>
                     <given-names>BB</given-names>
                  </name>
               </person-group>
               <article-title>Localizing the sentinel node in cutaneous melanoma: gamma probe detection versus blue dye</article-title>
               <source>Ann Surg Oncol</source>
               <year>1997</year>
               <volume>4</volume>
               <fpage>156</fpage>
               <lpage>60</lpage>
               <pub-id pub-id-type="pmid">9084853</pub-id>
               <pub-id pub-id-type="doi">10.1007/BF02303799</pub-id>
            </citation>
         </ref>
         <ref id="B38">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Messina</surname>
                     <given-names>JL</given-names>
                  </name>
                  <name>
                     <surname>Glass</surname>
                     <given-names>LF</given-names>
                  </name>
                  <name>
                     <surname>Cruse</surname>
                     <given-names>CW</given-names>
                  </name>
                  <name>
                     <surname>Berman</surname>
                     <given-names>C</given-names>
                  </name>
                  <name>
                     <surname>Ku</surname>
                     <given-names>NN</given-names>
                  </name>
                  <name>
                     <surname>Reintgen</surname>
                     <given-names>DS</given-names>
                  </name>
               </person-group>
               <article-title>Pathologic examination of the sentinel lymph node in malignant melanoma</article-title>
               <source>Am J Surg Pathol</source>
               <year>1999</year>
               <volume>23</volume>
               <fpage>686</fpage>
               <lpage>90</lpage>
               <pub-id pub-id-type="pmid">10366151</pub-id>
               <pub-id pub-id-type="doi">10.1097/00000478-199906000-00008</pub-id>
            </citation>
         </ref>
         <ref id="B39">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Yan</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Brennick</surname>
                     <given-names>JB</given-names>
                  </name>
               </person-group>
               <article-title>False-positive rate of the immunoperoxidase stains for MART1/MelanA in lymph nodes</article-title>
               <source>Am J Surg Pathol</source>
               <year>2004</year>
               <volume>28</volume>
               <fpage>596</fpage>
               <lpage>600</lpage>
               <pub-id pub-id-type="pmid">15105646</pub-id>
            </citation>
         </ref>
         <ref id="B40">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Mahmood</surname>
                     <given-names>MN</given-names>
                  </name>
                  <name>
                     <surname>Lee</surname>
                     <given-names>MW</given-names>
                  </name>
                  <name>
                     <surname>Linden</surname>
                     <given-names>MD</given-names>
                  </name>
                  <name>
                     <surname>Nathanson</surname>
                     <given-names>SD</given-names>
                  </name>
                  <name>
                     <surname>Hornyak</surname>
                     <given-names>TJ</given-names>
                  </name>
                  <name>
                     <surname>Zarbo</surname>
                     <given-names>RJ</given-names>
                  </name>
               </person-group>
               <article-title>Diagnostic Value of HMB-45 and Anti-Melan A Staining of Sentinel Lymph Nodes with Isolated Positive Cells</article-title>
               <source>Mod Pathol</source>
               <year>2002</year>
               <volume>15</volume>
               <fpage>1288</fpage>
               <lpage>93</lpage>
               <pub-id pub-id-type="pmid">12481009</pub-id>
               <pub-id pub-id-type="doi">10.1097/01.MP.0000037313.33138.DF</pub-id>
            </citation>
         </ref>
         <ref id="B41">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Shidham</surname>
                     <given-names>VB</given-names>
                  </name>
                  <name>
                     <surname>Susnik</surname>
                     <given-names>B</given-names>
                  </name>
                  <name>
                     <surname>Rao</surname>
                     <given-names>RN</given-names>
                  </name>
                  <name>
                     <surname>Chivukula</surname>
                     <given-names>M</given-names>
                  </name>
                  <name>
                     <surname>Kampalath</surname>
                     <given-names>B</given-names>
                  </name>
                  <name>
                     <surname>Komorowski</surname>
                     <given-names>R</given-names>
                  </name>
               </person-group>
               <article-title>HMB45 immunoreactive single cells in sentinel lymph nodes negative for melanoma micrometastases are mast cells</article-title>
               <source>Mod Pathol</source>
               <year>2004</year>
               <volume>17</volume>
               <fpage>1a</fpage>
               <lpage>388a</lpage>
               <pub-id pub-id-type="pmid">14702091</pub-id>
            </citation>
         </ref>
         <ref id="B42">
            <citation citation-type="journal">
               <person-group person-group-type="author">
                  <name>
                     <surname>Shidham</surname>
                     <given-names>VB</given-names>
                  </name>
                  <name>
                     <surname>Qi</surname>
                     <given-names>DY</given-names>
                  </name>
                  <name>
                     <surname>Acker</surname>
                     <given-names>S</given-names>
                  </name>
                  <name>
                     <surname>Kampalath</surname>
                     <given-names>B</given-names>
                  </name>
                  <name>
                     <surname>Chang</surname>
                     <given-names>CC</given-names>
                  </name>
                  <name>
                     <surname>George</surname>
                     <given-names>V</given-names>
                  </name>
                  <name>
                     <surname>Komorowski</surname>
                     <given-names>R</given-names>
                  </name>
               </person-group>
               <article-title>Evaluation of micrometastases in sentinel lymph nodes of cutaneous melanoma: Higher diagnostic accuracy with Melan-A and MART-1 compared to S-100 protein and HMB-45</article-title>
               <source>Am J Surg Pathol</source>
               <year>2001</year>
               <volume>25</volume>
               <fpage>1039</fpage>
               <lpage>46</lpage>
               <pub-id pub-id-type="pmid">11474288</pub-id>
               <pub-id pub-id-type="doi">10.1097/00000478-200108000-00008</pub-id>
            </citation>
         </ref>
      </ref-list>
   </back>
</article>
