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. 2011 Jul 15;124(14):2401–2413. doi: 10.1242/jcs.083782

Fig. 7.

Fig. 7.

Intracellular trafficking of FFT and FTT. HeLa cells were transfected with FLAG-tagged (A) FFT or (B) FTT construct for 24 hours and incubated with anti-FLAG antibodies for 45 minutes on ice followed by washes with PBS. Monolayers were either fixed and permeabilised (0 min) or incubated in serum-free media for 30, 60 minutes or 120 minutes at 37°C and then fixed and permeabilised. Monolayers were stained for the internalised antibody-bound FLAG–FFT and FLAG–FTT with Alexa-Fluor-568-conjugated anti-rabbit IgG (red) and (A) for GM130 using mouse monoclonal anti-GM130 antibodies (green) or LAMP1 using mouse monoclonal anti-LAMP1 antibodies (green) and (B) for GM130 using mouse monoclonal anti-GM130 antibodies (green) or CD63 using mouse monoclonal anti-CD63 antibodies (green). (C) Quantification of FLAG–FFT and FLAG–FTT levels within the Golgi. The percentage (±s.e.m.) of the total FLAG pixels that overlapped with GM130 in each cell after 60 minutes internalisation was determined using the plug-in OBCOL on ImageJ (n=20 for each sample). The data from FLAG–FFT and FLAG–FTT is expressed as a percentage of the FLAG–furin data set. ***P<0.001. Scale bars: 10 μm.