Human Cortical Neuronal Cells (HCN-1A Line)

Immunofluorescence with mouse anti-alpha-tubulin was employed to visualize distribution of the microtubule network in the human cortical neuronal cell culture illustrated above. The secondary antibody (goat anti-mouse IgG) was conjugated to Cy2 and mixed with Alexa Fluor 568 conjugated to phalloidin to simultaneously image tubulin and the actin cytoskeleton. Nuclei were counterstained with Hoechst 33258. Images were recorded in grayscale with a 12-bit digital camera coupled to either a Nikon E-600 or Eclipse 80i microscope equipped with bandpass emission fluorescence filter optical blocks. During the processing stage, individual image channels were pseudocolored with RGB values corresponding to each of the fluorophore emission spectral profiles.