Isolating Fluorescently Labeled Neurons from a Murine Brain
Trascrizione
Take a transgenic mouse cortex containing fluorescent protein-expressing neurons.
Secure it on a sample holder.
Mount the holder on the vibratome tray containing oxygenated aCSF to maintain the tissue's physiological conditions.
Using set parameters, obtain coronal slices.
Place the slices in a meshed holder containing oxygenated aCSF and activity blockers to stabilize them.
Treat the slices with proteases to digest the extracellular matrix and loosen cells.
Wash with aCSF to remove the proteases.
Transfer the slices to a culture plate with aCSF and serum.
Under a fluorescence microscope, dissect slice regions containing fluorescent neurons.
Transfer the dissected fragments to a tube containing aCSF and serum.
Mechanically dissociate the tissue to form a single-cell suspension. Transfer the cells to a culture plate containing aCSF.
Under a fluorescence microscope, use a capillary pipette to collect fluorescent neurons.
Dispense these neurons into a culture plate containing aCSF for further analysis.
