Semi-quantitative RT-PCR analysis of oncogene and tumor suppressor genes. A: A panel of 12 putative oncogenes and 6 TSG were analyzed, with the relative levels of mRNA expression as follows: negative band (-), faintly visible band (+/-), visible band (+), strongly visible band (++), N = Not Done. β-actin served as the internal comparative control. The grey values of PCR products of each gene are analyzed by the AlphaEase 3 software and standardized according to β-actin in every sample. B: a, b, Daughter melanoma cell lines secrete SPP1 (osteopontin). The melanoma cell lysates and conditioned cell-free media was resolved by SDS-PAGE and transferred to a PVDF membrane. The blot was probed with anti-SPP1 antibody. c-f, Antibodies for DSC-3 (c), CLCA2 (d), PDGFRL (e) and α-tubulin (f) as an internal control. Lanes 1–3 are PCM and lanes 4–6 are MM cell lines.