Autophagy assay cell type - RIN-5F

As autophagy is a multi-step process which includes not just the formation of autophagosomes, but most importantly, flux through the entire system, including the degradation upon fusion with lysosomes, which makes it quite challenging for detection. There are several methods for detection in mammalian cells, including immunoblotting analysis of LC3 and p62 and detection of autophagosome formation/maturation by fluorescence microscopy, Currently, there is no single “gold standard” for determining the autophagic activity that is applicable in every experimental context, hence it is recommended to go for the combined use of multiple methods to accurately assess the autophagic activity in any given biological setting.

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Found 1 matching solution for this experiment

LC3A/B (D3U4C) XP® Rabbit mAb

Cell Signaling Technology

Upstream tips	Protocol tips	Downstream tips
- Lyse cells in Laemmli buffer containing 60 mM Tris-HCl pH 6.8, 2% (w/v) SDS, 10 mM EDTA, 10% (w/v) glycerol	- Dilute Primary Ab at -1:1000. - Incubate Ab for overnight at 4°C

Upstream tips
- Lyse cells in Laemmli buffer containing 60 mM Tris-HCl pH 6.8, 2% (w/v) SDS, 10 mM EDTA, 10% (w/v) glycerol
Protocol tips
- Dilute Primary Ab at -1:1000. - Incubate Ab for overnight at 4°C

Manufacturer protocol Publication protocol 1 Relevant paper

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