Analysis of SPI-6 expression in stem cell lines. (A) Protein extracts of maGSC Stra8 SSC5 cells cultured in basic medium (condition I), medium plus LIF (condition II), basic medium on MEFs (condition III), or in medium plus LIF on MEFs (condition IV) were separated by SDS-PAGE and the blot was probed with an anti-SPI-6 Ab and with an anti-HSC70 mAb as loading control. A protein extract of activated CTLs from OT-I mice served as positive control for SPI-6 detection. (B) The expression of SPI-6 was similarly probed in differentiated cells derived from maGSC Stra8 SSC5 cells and in two ESC lines for comparison. (C) Expression of Serpinb9 gene that encodes SPI-6, was analyzed in triplicates by qRT-PCR in the same cell lines in comparison to the housekeeping gene Hprt1. The maGSC Stra8 SSC5 cells had been cultured under conditions I, II, III, and IV before mRNA preparation. Means and SDs of the Δct values (Hprt minus gene of interest) are shown.