Cells were
fixed for 20 min at room temperature with 4% formaldehyde, permeabilized in 0.5% Triton X-100 and stained with primary antibodies (anti-mouse FLAG and anti-rabbit LAMP 1) for 30 min at
room temperature, washed and stained with Alexa Fluor 488-, or
549-nm-coupled secondary antibodies and mounted as described
[3]. Slides were dried, stored in the dark, and analyzed the next day
by confocal microscopy analysis using a Zeiss LSM-710 META microscope (Carl Zeiss).