Fig. 1.

Workflow to generate normal mesothelial cultures. On day 0, patient pleural sample is minced, placed in an etched culture dish, and incubated to allow for attachment of tissue fragments to the plate bottom. By day 14 (explant culturing), cells have migrated out onto the plate surface and are ready for harvest. Black arrows indicate direction of growth leaving the tissue fragment. Dashed line indicates boundary of tissue. Live-cell FACS recovers human mesothelial cell (HMC) according to CDH1 and CD71 surface expression. At day 16 cells are transduced with human telomerase reverse transcriptase (hTERT)-neoR lentivirus. Cells are exposed to neomycin over 5 days to ensure effective selection by resistance. By day 23, normal HMC are ready for propagation in vitro.