and were loosened by pipetting after washing several times with pbs the cells were suspended in dmem containing 10 fcs and were cultivated on a 10 cm dish to investigate the localization of endogenous ys68 within cells cultured hepatic cells were stained with anti ys68 antibodies first the cells were fixed with 4 formalin and then treated with 0 1 triton x 100 for cell staining next cells were reacted with the primary antibodies and then with secondary antibodies the cells were visualized in the same manner as in example 5 consequently although ys68 has multiple nuclear transport signals strong expression was found not only in the nucleus but also around the nucleus which expression depended on cells fig1 next similar analysis for the expression in hematocytes was carried out ys68 expression in hematocytes separated from embryonic liver was found to have varied strengths of expression depending on the cell type fig1 b therefore the group of hematocytes was sorted using cd34 which is a marker for immature hematocytes and ys68 expression in cd34 positive cells was investigated to collect cd34 positive cells embryonic liver e14 5 was incubated in a dissociation buffer at 37 u00b0 c for 30 minutes and then the cells were dissociated by pipetting in pbs after passing through a nylon mesh filter falcon the cells were suspended in a sample buffer 0 5 bsa 2 mm edta in pbs the cells were reacted with biotin labeled anti cd34 antibodies pharmingen followed by fitc labeled streptavidin at 4 u00b0 c and then were incubated with anti fitc microbeads cd34 positive cells were eluted using macs magnetic cell sorting column according to the instructed protocol the cells were centrifuged on a slide glass at 400 rpm for 5 minutes to.