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Fig. 4. (A) Growth rates of Kit^+/+ and Kit^W-lacZ/W-FKB cells in the presence and absence of AP20187 during routine passage, expressed as the total number of cells present 2 days after plating 1x10⁶ cells. The solid bars within the individual scatters represents the median of six independent passages. (B) Total numbers of stem cell (SC), differentiated (DIFF) or mixed (MIX) colonies produced from Kit^+/+ (+/+), Kit^W-lacZ/+ (+/-), Kit^{W-lacZ/W-lacZ} (-/-) and Kit^W-lacZ/W-FKB (-/FKBKit) in 100 or 0 U/ml of LIF in the presence of AP20187. The results shown are representative examples of three assays carried out in duplicate. (C) Combined number of mixed and differentiated colonies produced from Kit^+/+ (+/+), Kit^W-lacZ/+ (+/-), Kit^{W-lacZ/W-lacZ} (-/-) and Kit^W-lacZ/W-FKB (-/FKB) ES cells 5 days after plating at low density in 100 U/ml LIF and 10 nM AP20187. The graphs show the mean (± s.e.m.) calculated from three independent experiments carried out in duplicate. (D) Western blot analysis of protein lysates isolated from Kit^+/+, Kit^W-lacZ/+, Kit^{W-lacZ/W-lacZ} and Kit^W-lacZ/W-FKB ES cells grown in the presence (+) or absence (-) of AP20187 as indicated. Blots were sequentially probed with an antibody specific to KIT phosphotyrosine 730 (pY730) then an anti-GAPDH (GAPDH) antibody as a loading control.

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