2.4.2. Fluorescent labelling of living neurons
Transfected neurons were washed with Neurobasal media three times. Then,
live-cell labelling experiments were conducted with microtubule-specific
and lysosomal specific fluorescence dye, respectively. The
tubulin-tracker dye was diluted with Neurobasal media (1:500) and 200 μL
was added to the coverslip. The lyso-tracker dye was diluted with
Neurobasal media (1:1000) and added to the coverslip. Neurons were
labelled in 37℃, 5% CO2 cell incubator for 30 min.
After 3 times washed with PBS, 100 μL Hoechst solution was used for
nucleus staining. After PBS washing, coverslips were immersed by 1 mL
Neurobasal media in the 3 cm diameter dish and observed under an upright
microscope. In the experiment, Neurobasal media, PBS, and Hoechst
solution were preheated in a water bath at 37℃ to maintain the normal
physiological status of neurons.