Autophagy assay cell type - Goblet cells

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 4 matching solutions for this experiment

SQSTM1 Antibody (D-3)

Santa Cruz Biotechnology

Upstream tips	Protocol tips	Downstream tips
Lyse cells in RIPA buffer (PBS, pH 7.4, 1.0% IPEGAL CA630; 0.5% sodium deoxycholate; 0.1% SDS containing freshly added proteinase inhibitors	Primary antibody dilution 1:1000

Upstream tips
Lyse cells in RIPA buffer (PBS, pH 7.4, 1.0% IPEGAL CA630; 0.5% sodium deoxycholate; 0.1% SDS containing freshly added proteinase inhibitors
Protocol tips
Primary antibody dilution 1:1000

Manufacturer protocol Publication protocol 1 Relevant paper

Anti-LC3B antibody produced in rabbit

Sigma-Aldrich

Upstream tips	Protocol tips	Downstream tips
Lyse cells in RIPA buffer (PBS, pH 7.4, 1.0% IPEGAL CA630; 0.5% sodium deoxycholate; 0.1% SDS containing freshly added proteinase inhibitors	Primary antibody dilution 1:750

Upstream tips
Lyse cells in RIPA buffer (PBS, pH 7.4, 1.0% IPEGAL CA630; 0.5% sodium deoxycholate; 0.1% SDS containing freshly added proteinase inhibitors
Protocol tips
Primary antibody dilution 1:750

Manufacturer protocol Publication protocol 1 Relevant paper

LC3B Antibody Kit for Autophagy

Thermo Fisher Scientific

Upstream tips	Protocol tips	Downstream tips
Antibody detects two bands corresponding to LC3b-I and LC3b-II.	1:1500 dilution of primary antibody

Upstream tips
Antibody detects two bands corresponding to LC3b-I and LC3b-II.
Protocol tips
1:1500 dilution of primary antibody

Manufacturer protocol Publication protocol 1 Relevant paper

SQSTM1 Antibody (D-3)

Santa Cruz Biotechnology

Upstream tips	Protocol tips	Downstream tips
	Primary antibody dilution 1:1000

Protocol tips
Primary antibody dilution 1:1000

Manufacturer protocol Publication protocol 1 Relevant paper

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