Autophagy assay cell type - HaCaT

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 3 matching solutions for this experiment

LC3A/B Antibody #4108

Cell Signaling Technology

Upstream tips	Protocol tips	Downstream tips
LC3A/B Antibody detects endogenous levels of total LC3A and LC3B proteins. Cross-reactivity may exist with LC3C. Stronger reactivity is observed with the type II form of LC3A/B.	Dilute primary Ab at 1:1000 and incubate overnight at 4℃ and for IF at 1:200

Upstream tips
LC3A/B Antibody detects endogenous levels of total LC3A and LC3B proteins. Cross-reactivity may exist with LC3C. Stronger reactivity is observed with the type II form of LC3A/B.
Protocol tips
Dilute primary Ab at 1:1000 and incubate overnight at 4℃ and for IF at 1:200

Manufacturer protocol Publication protocol 1 Relevant paper

Beclin-1 (D40C5) Rabbit mAb

Cell Signaling Technology

Upstream tips	Protocol tips	Downstream tips
Beclin-1 (D40C5) Rabbit mAb detects endogenous levels of total Beclin-1 protein."	Dilute primary Ab at 1:1000 and incubate overnight at 4℃ and for IF at 1:200

Upstream tips
Beclin-1 (D40C5) Rabbit mAb detects endogenous levels of total Beclin-1 protein."
Protocol tips
Dilute primary Ab at 1:1000 and incubate overnight at 4℃ and for IF at 1:200

Manufacturer protocol Publication protocol 1 Relevant paper

Atg7 (D12B11) Rabbit mAb

Cell Signaling Technology

Upstream tips	Protocol tips	Downstream tips
Atg7 (D12B11) Rabbit mAb recognizes endogenous levels of total Atg7 protein.	Dilute primary Ab at 1:1000 and incubate overnight at 4℃ and for IF at 1:200

Upstream tips
Atg7 (D12B11) Rabbit mAb recognizes endogenous levels of total Atg7 protein.
Protocol tips
Dilute primary Ab at 1:1000 and incubate overnight at 4℃ and for IF at 1:200

Manufacturer protocol Publication protocol 1 Relevant paper

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