Figure 2.

Expression of nestin, CD133 and other intracellular proteins in the osteosarcoma cell lines. Representative expression of vimentin, desmin, and S100 protein in the osteosarcoma cell lines: vimentin expression in OSA-03 cell line (a); desmin expression in OSA-02 cell line (b); S100 protein expression in OSA-01 cell line (c). Medium or strong expression of nestin was found in three newly derived cell lines and the distinct network of nestin-positive filaments was observable in the OSA-03 cell line (h, i). OSA-05 cell line was nestin-negative (k, l). Expression of CD133 cell surface molecule (g, i, j, l) was found in all osteosarcoma cell lines and co-expression of nestin and CD133 was confirmed using double labeling in nestin-positive cell lines (g-i). Representative double labeling for CD133 and nestin in nestin-positive OSA-03 (g-i) and nestin-negative OSA-05 (j-l) newly derived osteosarcoma cell lines. Glioblastoma GM-7 cell line labeled by the same antibodies was used as positive control (d) and control established Saos-2 osteosarcoma cell line also showed strong positivity for both nestin (e) and CD133 (f). Vimentin (a, red), S100 protein (c, red), and CD133 (d, f, g, i, j, l, red) stained by indirect immunofluorescence using TRITC-labeled secondary antibody; desmin (b, green) and nestin (d, e, h, i, k, l, green) stained by the same method using FITC-labeled secondary antibody; counterstaining with DAPI (a-f, i, l). Bars, 30 μm (a-c), 20 μm (d), 15 μm (e-l).