Quantifying autophagosomes and autolysosomes in cells using imaging flow cytometry.

Cytometry. Part a : the Journal of the International Society for Analytical Cytology
Robin RajanNinh M La-Beck

Abstract

Autophagy dysregulation has been implicated in numerous diseases and many therapeutic agents are known to modulate this pathway. Therefore, the ability to accurately monitor autophagy is critical to understanding its role in the pathogenesis and treatment of many diseases. Recently an imaging flow cytometry method measuring colocalization of microtubule associated protein 1B light chain 3 (LC3) and lysosomal signals via Bright Detail Similarity (BDS) was proposed which enabled evaluation of autophagic processing. However, since BDS only evaluates colocalization of LC3 and lysosomal signals, the number of autophagy organelles was not taken into account. We found that in cells classified as having Low BDS, there was a large degree of variability in accumulation of autophagosomes. Therefore, we developed a new approach wherein BDS was combined with number of LC3+ puncta, which enabled us to distinguish between cells having very few autophagy organelles versus cells with accumulation of autophagosomes or autolysosomes. Using this method, we were able to distinguish and quantify autophagosomes and autolysosomes in breast cancer cells cultured under basal conditions, with inhibition of autophagy using chloroquine, and with induction ...Continue Reading

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Citations

Nov 10, 2015·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Tobias BraunAgnes Csiszár
Sep 14, 2016·Methods : a Companion to Methods in Enzymology·Brian K McFarlin, Melody A Gary
Jan 21, 2017·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Attila Tárnok
Jun 18, 2019·Cytometry. Part a : the Journal of the International Society for Analytical Cytology·Jessica P Houston
Mar 7, 2021·Cell Death & Disease·Carolyn N BrownCharles L Edelstein

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