Atg3 Antibody

Protocol tips

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	Dilute primary Ab at 1:1,000 and incubate overnight at 4C.

Protocol tips
Dilute primary Ab at 1:1,000 and incubate overnight at 4C.

Publication protocol

Total protein (30 μg) and pre-stained molecular weight markers were separated by SDS-PAGE and electro-transferred onto a polypropylene difluoride membrane (Millipore, Billerica, MA, USA). The membranes were blocked by incubating with Tris-buffered saline containing 0.1% Tween-20 and 5% non-fat milk for 2 h at room temperature. Then, the membranes were probed at 4°C overnight with the following primary antibodies: from Cell signaling (Danvers, MA, USA): rabbit anti-Atg3 (catalog no.:3415; 1:1000 dilution), rabbit anti-LC3B (12741; 1:1000), rabbit anti-GAPDH (5174; 1:1000), rabbit anti-p62 (8025; 1:1000), rabbit anti-p-AKT (S473) (9271; 1:1000), rabbit anti-p-mTOR (S2448) (5536; 1:1000), rabbit anti-caspase-3 (9662; 1:1000) and mouse anti-caspase-8 (9746; 1:1000) antibodies; from Zen BioScience (Chengdu, Sichuan, China): mouse anti-FLAG (390002; 1:2000). After washing, the membrane was incubated with a secondary antibody conjugated to horseradish peroxidase for 1 h at room temperature. Finally, the membrane was prepared with enhanced chemiluminescence substrate (Pierce, Rockford, IL, USA) and visualized on a LAS-3000 Luminescence Image Analyzer (Fujifilm, Tokyo, Japan). All antibodies were purchased from Cell Signaling Tech Inc.

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Papers

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Manufacturer protocol

Download the product protocol from Cell Signaling Technology for Atg3 Antibody below.

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Videos

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