Figure 1. Differential endogenous OPG status of eighteen human melanoma cell lines.

A) Some melanoma cell lines constitutively produce soluble OPG. Soluble OPG production by human melanoma cells and control (prostate and embryonic kidney) cells after 72 h culture was assessed by specific ELISA. Data are mean ±standard deviation of five replicates in a single experiment; one of 4 representative experiments shown. B) Intracellular staining for OPG correlates with production of soluble OPG. Open histograms, unstained; shaded histograms, stained with specific mAb. Staining with isotype control was not different than unstained (not shown). Data are representative intracellular staining for OPG in four melanoma cell lines, assessed after 72h in culture. Intracellular OPG was detected (>5% positive events in the tumor gate) in all samples that tested positive for OPG by ELISA (> 31pg/ml), and intracellular OPG was absent from cells that were negative for soluble OPG (<31pg/ml). Data from one of three similar experiments. C) OPG gene expression correlates with production of soluble OPG. Expression of OPG genetic message was assessed in 18 human melanoma cell lines, relative to the housekeeping gene GADPH, by real-time PCR. Data are mean ±standard deviation of three replicate measures in one of two representative experiments.