title: Novel application of a fibrin cell block method to evaluate melanocytic cell populations.
abstract: Confirming melanocytic lineage and purity is important for experiments using cultured human melanocytes. The objective of this study was to develop a simple, reliable method to evaluate and archive cultured melanocytic cells. Melanocytes were isolated from adult skin biopsies or from neonatal foreskins using standard culturing methods. Fibrin cell blocks (FCBs) were prepared from cultured cells at passages two and six. Fibrin blocks were paraffin-embedded and sectioned for immunohistochemical (CD68, Melan-A, and HMB-45) and H & E staining. Flow cytometry was performed (Melan-A) at passage six. A mixing experiment with cultured melanocytes and fibroblasts was performed and cell population purity was determined by manual counts of positively staining cells in the FCBs and by flow cytometry. The FCB method of evaluating population purity was validated experimentally and by correlation with flow cytometry results. Preparation of a FCB followed by immunohistochemical staining is an easy and inexpensive way to confirm melanocytic lineage, estimate population purity, and provide a permanent archive of cultured cells.
authors: Department of Dermatology, University of Utah Health Sciences Center, Salt Lake City, Utah 84132, USA. scott@derm.med.utah.edu
authors: Scott R
authors: SR
authors: Florell
authors: Soo J
authors: SJ
authors: Schmidt
authors: Patricia
authors: P
authors: Porter-Gill
authors: Kurt H
authors: KH
authors: Albertine
authors: Kelley J
authors: KJ
authors: Murphy
authors: Courtney B
authors: CB
authors: McKinney
authors: Kenneth M
authors: KM
authors: Boucher
authors: Douglas
authors: D
authors: Grossman
authors: Diana L
authors: DL
authors: Biddle
authors: Frederic
authors: F
authors: Clayton
authors: Lester J
authors: LJ
authors: Layfield
authors: Sancy A
authors: SA
authors: Leachman
mesh_terms: D000328:Adult; D015703:Antigens, CD / Q000276:immunology; D015214:Antigens, Differentiation, Myelomonocytic / Q000276:immunology; D000951:Antigens, Neoplasm; D002478:Cells, Cultured; D005337:Fibrin / Q000737:chemistry*; D005347:Fibroblasts / Q000166:cytology*; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D058965:MART-1 Antigen; D008544:Melanocytes / Q000166:cytology*; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins / Q000276:immunology; D012867:Skin / Q000166:cytology*; D000097382:CD68 Molecule
chemical_list: D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D000951:Antigens, Neoplasm; D005337:Fibrin; D058965:MART-1 Antigen; D058950:Melanoma-Specific Antigens; D009363:Neoplasm Proteins; C067980:CD68 antigen, human; D000097382:CD68 Molecule; C548667:MLANA protein, human