HS4C3-Binding Epitope Contributes to Fas Signaling Figure 1. Increase in the HS4C3-binding epitope induces differentiation. (A) FACS analysis, using the HS4C3 antibody, of mESCs after LIF withdrawal up to day 8. mESCs were cultured in the presence of LIF until day 0. In the left panel, a histogram shows a representative result of the FACS analysis (black line, in the presence of LIF; red line, 4 days after LIF withdrawal; green line, 6 days after LIF withdrawal; blue line, 8 days after LIF withdrawal). The gray line shows the result obtained from cells not treated with primary antibody. In the right panel, the values shown are the mean fluorescence intensity 6 SD after normalization against mESCs cultured in the presence of LIF (arbitrary value = 1). (B) Immunostaining, using the HS4C3 antibody, of non-permeabilized mESCs cultured in the presence of LIF (+LIF) or mESCs cultured for 7 days in the absence of LIF (2LIF) (upper panel). Lower panel shows DIC images. Scale bar, 5 mm. Representative confocal images are shown. (C) Real time PCR analysis of mESCs after LIF PLOS ONE | www.plosone.org 3 August 2012 | Volume 7 | Issue 8 | e43440