A new live-cell biobank workflow efficiently recovers heterogeneous melanoma cells from native biopsies.
Fibroblast contamination can make establishing primary melanoma cell cultures from native biopsies a major challenge, due to fibroblasts overgrowing the melanoma cells. Standard protocols therefore enrich for highly proliferative melanoma cells that grow well in vitro but may not represent the full range of in vivo tumor heterogeneity. Here we apply conditional methods that more effectively retrieve melanoma cells by differential trypsinization or by inducing fibroblast senescence through contact inhibition, serum starvation or deprivation of adhesion. Simple mixing experiments of melanoma and fibroblast cells demonstrated the efficacy of the new protocols in retrieving slow-growing melanoma cells. Applying our protocols to 20 cultures that had failed to grow by conventional methods, we could retrieve 12 (60%) validated melanoma cell cultures. Further application of the protocols in the live-cell biobank of 124 early passage cultures significantly improved recovery rates from 13% using standard protocols to 70% overall for the new workflow.
Department of Dermatology, University Hospital Zürich and University of Zurich, Zürich, Switzerland.
Marieke I G
MI
Raaijmakers
Daniel S
DS
Widmer
Melanie
M
Maudrich
Tabea
T
Koch
Alice
A
Langer
Anna
A
Flace
Claudia
C
Schnyder
Reinhard
R
Dummer
Mitchell P
MP
Levesque
D018070:Biological Specimen Banks; D001706:Biopsy; D002469:Cell Separation / Q000379:methods; D005347:Fibroblasts / Q000473:pathology; D006801:Humans; D008545:Melanoma / Q000235:genetics / Q000473:pathology* / Q000556:secondary; D009154:Mutation; D061251:Primary Cell Culture / Q000379:methods*; D012878:Skin Neoplasms / Q000235:genetics / Q000473:pathology*; D014407:Tumor Cells, Cultured; D057188:Workflow
biobank; cell culture; fibroblast overgrowth; melanoma