Immunofluorescence detection of synemin in mouse ES cells. A, Western blotting showing the specificity of synemin antibodies with proteins extracted from mouse bladder. The antibody anti-synemin H detected one band of 180 kDa, corresponding to synemin H isoform (lanes 1-2), and the anti-synemin H/M detected two bands 150 kDa and 180 kDa that correspond to H and M isoforms (lanes 3-4). GAPDH was used for the loading control (lanes 5-6). B, Diagram illustrating the regions recognized by antibodies anti-synemin H and anti-synemins H/M. C-K, Immunochemistry analysis demonstrating ES cells colonies labeled for Oct-3/4 (C), keratin 8 (F), SSEA-1 (I), synemin M (D, G) and synemin H (J). Synemin M was present with Oct-3/4 (C-E) and keratin 8 (F-H) in pluripotent ES cells. No immunolabelling for synemin H was observed in undifferentiated ES cells expressing SSEA-1 (I-K). Syn: synemin; K8: keratin 8. Bar = 20 μm.