Autophagy assay cell type - MMQ

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.

Start discussion

No discussions found

Share your thoughts or question with experts in your field

Start a discussion

Found 3 matching solutions for this experiment

Anti-LC3B antibody produced in rabbit

Sigma-Aldrich

Upstream tips	Protocol tips	Downstream tips
- RIPA buffer is composed of 1 × PBS, 1% Nonidet P-40, 0.5% sodium deoxycholate, 0.1% SDS, 1 mM phenyl methylsulfonyl fluoride, and protease inhibitors for better results

Upstream tips
- RIPA buffer is composed of 1 × PBS, 1% Nonidet P-40, 0.5% sodium deoxycholate, 0.1% SDS, 1 mM phenyl methylsulfonyl fluoride, and protease inhibitors for better results

Manufacturer protocol Publication protocol 1 Relevant paper

Anti-SQSTM1 / p62 antibody

Abcam

Upstream tips	Protocol tips	Downstream tips
	- Dilute primary Ab in blocking buffer with 0.1% saponin. - Incubate cells at 4°C overnight

Protocol tips
- Dilute primary Ab in blocking buffer with 0.1% saponin. - Incubate cells at 4°C overnight

Manufacturer protocol Publication protocol 1 Relevant paper

Beclin-1 Antibody

Cell Signaling Technology

Upstream tips	Protocol tips	Downstream tips
	- Dilute primary ab 1:1000 and incubate at overnight at 4°C

Protocol tips
- Dilute primary ab 1:1000 and incubate at overnight at 4°C

Manufacturer protocol Publication protocol 1 Relevant paper

Can't find the product you've used to perform this experiment? It would be great if you can help us by Adding a product!

Fill out your contact details and receive price quotes in your Inbox