Crowd4Discovery autophagy experiment 1
Ethan Perlstein, Daniel Korostyshevsky
In this pilot experiment, Danny (@badomens) has treated fixed cultured ventral midbrain neurons with a pH-sensitive dye called pHrodo, which accumulates and "glows" red inside acidic compartments of the endocytic pathway. Danny is comparing untreated neurons ("steady-state") vs 24-hr amphetamine-treated neurons ("400µM").
He also treated these cells with an antibody that recognizes a protein called LC3, which marks intracellular structures called autophagosomes, which deliver cellular bits to the recycling centers of cells. This LC3-specific antibody was visualized with a second, fluorescent antibody that "glows" green. Yellow indicates co-localization of endocytic compartments with autophagosomes.
This experiment is now being done using a confocal microscope to thoroughly assess colocalization of the two labels.