Figure 12.

GFP fluorescence in the autolysosome can be recovered upon neutralization of the pH. (A) GFP-LC3 emits green fluorescence in the autolysosomes of post-mortem processed heart sections. Cryosections of 3.8% paraformaldehyde-fixed ventricular myocardium from 3-wk-old GFP-LC3 transgenic mice at the baseline (control) or starved for 24 h (starved) were processed for immunostaining using a standard protocol (buffered at pH 7.4). Most of the GFP-LC3 puncta are positive for LAMP1, suggesting that the autolysosomes had recovered GFP fluorescence. (B) Colocalization between GFP-LC3 direct fluorescence (green) and indirect immunostaining for GFP (red). Sections processed as in (A) were immunostained for GFP using a red fluorescence-tagged secondary antibody, and the colocalization with GFP fluorescence was examined by confocal microscopy. Almost all of the red puncta emit green fluorescence. Image provided by Xuejun Wang.