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Jan Klos

EQA of melanoma marker analyses

Handout (pdf, 32 p)

Handout (pdf, 8 p)


The most essential immunohistochemical markers useful in differential diagnosis of melanocytic lesions were tested numerous times.
Melanosome Specific Antigen: tested in 3 runs (total 323 participants). The only clone used was mAb HMB-45. The rate of sufficient staining above 90% in last two runs, and 60% optimal results during the last run, confirm high quality of this antibody. Protocols based on HIER and a non-biotin based detection systems gave the highest proportion of optimal results. Main causes of insufficient staining were omission/insufficient HIER and biotin based detection systems. Recommended control: angiomyolipoma.
Melanocyte differentiation antigen (Melan A/MART1): tested in 6 runs (total 682 participants). Clone A103 used by 188/198 labs (95%) in the last run, is recommended. Optimal results were also obtained with clone M2-7C10 alone or in cocktails. Optimal (40%) and good (29%) staining resulted in 69% sufficient rate. Main causes of insufficient results: use of detection system with low sensitivity, insufficient HIER and too low concentration of primary Ab. Recommended control: adrenal gland for protocols with clone A103 and non-biotin based detection systems, and angiomyolipoma for other protocols.
S-100: tested in 3 runs with total 369 participants. Optimal results were achieved only with concentrated pAb Z0311, by 36/200 (18%) of participants. Overall sufficient staining achieved by 64% of labs was slightly less than in previous runs. HIER and appropriate concentration of the successful primary Ab were prerequisites of optimal performance. Proteolysis or omission of HIER generally gave significantly inferior performance. Recommended control: appendix or tonsil.
Ki-67 assessed in 4 runs (total 495 participants) showed 89% (205/229) sufficient results. Recommended clones are MIB1, MM1, K2, 30-9 and SP6. Efficient HIER (preferable in alkaline buffer) is mandatory for optimal result. The RTU products showed superior pass rates as compared to in-house validated assays. Causes of insufficient staining: too low concentration of primary Ab, insufficient/excessive HIER and inadequate deparaffination. Recommended control: tonsil.
Vimentin: assessed in 2 runs (total 243 participants). During last run 122/164 (74%) labs used V9 clone and 83% (136/164) achieved sufficient results. Optimal staining: the highest proportion obtained with V9 clone, but was also achievable with clones Vim 3B4 and SP20. Causes of insufficient staining: too low concentration of primary Ab, proteolytic pretreatment or omission of HIER. Recommended control: tonsil.

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Dr. Jan Klos M.D. is a Senior Consultant and a Head of the Immunohistochemistry Section in the Department of Pathology, Stavanger University Hospital, Norway.  He was educated at Medical University of Warsaw (1968-1974), trained and practiced pathology in Warsaw, Poland (1975-1982). Since 1982 he has been practicing and teaching pathology abroad, working first at the University of Jos, Nigeria (1982-84), later at the Central Hospitals in Falun and Gavle as well as at University of Umeå, Sweden (1984-1998), and since 1998 in Stavanger, Norway. His professional interest is diagnostic pathology with the main focus on hematopathology, pathology of the breast and cytopathology. He is a co-author of more than 30 publications in pathology. He is also a member of Editorial Board of Polish Journal of Pathology. Since 2009 he is a member of the core group in NordiQC, representing there also Norwegian Society of Pathologists, and from 2015 he is appointed as a Chairman of NordiQC. He has been actively involved in teaching immunohistochemistry since 2000 as an invited lecturer on various international and national courses and meetings. During 2008-2013 he was also a co-organizer of annual NordiQC Workshops in Immunohistochemistry and from 2014 he is an organizer of the NordiQC Academy courses “Immunohistochemistry for Pathologists”.

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