Fig. 2.

Expression of murine mesothelin. a RT-PCR detection of mesothelin (M, 449 base pairs) and control GAPDH (G, 473 bp) RNA in Panc02 cells. b Mesothelin protein surface expression – cells were stained with rat anti-mouse mesothelin monoclonal antibody B35 (black line) or normal rat IgG (gray line) and Cy5-conjugated donkey anti-rat IgG and analyzed by flow cytometry. Top right panels show that Panc02 surface detection is reduced by trypsin treatment compared to scraping of the adherent cells into solution. Adherent Panc02 cells were harvested by treatment with Trypsin/EDTA cocktail or by scraping. c Because trypsin reduced mesothelin detection, all subsequent experiments on adherent cells were performed using chelating agents without trypsin. Mesothelin expression was detectable on mouse-mesothelin-transfected HEK, and several mouse cancer cell lines: AB12 mesothelioma, TRAMP-C3 prostate adenocarcinoma, the JWF2 keratinocyte squamous cell carcinoma, and NIH/3 T3 cells