Fig. 2. p25SS transport to post-Golgi membranes. (A) HeLa cells were transfected with myc-p23SS, HA-p24SS or myc-p25SS, as indicated. Then, cells were either permeabilized and the distribution of the indicated proteins was revealed by immunofluorescence using antibodies against myc or HA, or not permeabilized (4°C) and the corresponding antibodies were added to the living cells at 4°C prior to fixation. (B) Cells transfected as in A were incubated for 16 hours at 37°C with antibodies against myc or HA to detect the indicated proteins, and then the distribution of the endocytosed antibodies was revealed by immunofluorescence. (C) Cells were triply transfected with untagged p23, HA-p24 and myc-p25SS. Then, cells were incubated for 16 hours at 37°C with antibodies as in B except that p23 was labeled with anti-lumenal domain antibodies. The distribution of the endocytosed antibodies was analyzed as in B. Scale bar: 10 µm.

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