Immunostaining of HeLa Cells and Neurons

Immunostaining was performed as described previously^{55 (link)}. HeLa cells or primary neurons cultured on glass cover slips were fixed for 10 min at room temperature with 3.7% formaldehyde in phosphate-buffered saline (PBS) and then incubated consecutively with primary antibodies and Alexa Fluor 488- or Alexa Fluor 546-labeled secondary antibodies in PBS containing 0.5% Triton X-100. Nuclei were also stained with Hoechst 33258 (Wako, Tokyo, Japan) as indicated. The cells were finally covered with a drop of GEL/MOUNT (Biomeda, Hayward, CA) and observed with an Olympus IX-81 fluorescence microscope (Olympus, Tokyo, Japan) or LSM800 confocal microscope (Zeiss, Tokyo, Japan).