Beclin-1 (D40C5) Rabbit mAb

Autophagy assay cell type - K562 cells

Experiment
Autophagy assay cell type - K562 cells
Product
Beclin-1 (D40C5) Rabbit mAb from Cell Signaling Technology
Manufacturer
Cell Signaling Technology

Protocol tips

Upstream tips
Lyse cells in lysis buffer (50 mM Tris-HCl, pH 7.5, 1% Nonidet P-40, 1 mM phenylmethylsulfonyl fluoride and 1 mM NaF) containing complete protease inhibitor cocktail
Protocol tips
Dilute primary antibody at 1:1,000 dilutionand incubate at 25°C for 2 h

Publication protocol

Cells were washed with PBS and then incubated with lysis buffer (50 mM Tris-HCl, pH 7.5, 1% Nonidet P-40, 1 mM phenylmethylsulfonyl fluoride and 1 mM NaF) containing complete protease inhibitor cocktail (Roche Applied Science, Mannheim, Germany) for 1 h at 4°C. The resulting supernatants were collected for protein ßquantitation using a bicinchoninic acid kit (Pierce; Thermo Fisher Scientific, Inc.). Crude proteins (20 µg) were subjected to 12.5% SDS-PAGE, transferred to a nitrocellulose membrane (EMD Millipore, Billerica, MA, USA) and then incubated with 5% w/v skimmed milk/PBS at room temperature for 1 h to block nonspecific binding. The blocked membrane was then incubated with primary antibodies (1:1,000 dilution) at 25°C for 2 h, followed by incubation with peroxidase-conjugated anti-IgG secondary antibodies (1:2,000 dilution) at 25°C for 1 h. Detailed information for antibodies were described in the reagents. The blocked membrane without incubation with primary antibodies was also incubated with secondary antibodies for specificity test, and a nonsignificant signal was detected as the control. Signal development was performed using an enhanced chemiluminescence reagent (EMD Millipore). Luminescent signals were acquired and quantified using an image analysis system (LAS-4000 with Image Reader LAS-4000 version 2.1, Fuji, Tokyo, Japan). GAPDH was used as an internal control.

Full paper   Login or join for free to view the full paper.

Reviews

Beclin-1 (D40C5) Rabbit mAb from Cell Signaling Technology has not yet been reviewed for this experiment

We'd love it if you would be the first to write a review!

Discussion

Start your discussion

Share your thoughts or question with experts in your field

Start a discussion

Papers

Check out relevant papers found by Labettor's AI that are relevant for performing Autophagy assay cell type - K562 cells using Beclin-1 (D40C5) Rabbit mAb from Cell Signaling Technology.

View full paper   Login or join for free to view the full paper.

Manufacturer protocol

Download the product protocol from Cell Signaling Technology for Beclin-1 (D40C5) Rabbit mAb below.

Download PDF Download manufacturer protocol

Videos

Check out videos that might be relevant for performing Autophagy assay cell type - K562 cells using Beclin-1 (D40C5) Rabbit mAb from Cell Signaling Technology. Please note that these videos are representative and steps or experiment specific processes must be kept in mind to expect desired results.

We haven't found any additional videos for this experiment / product combination yet.

Outsource your experiment

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

  Outsource experiment
Become shareholder Discussions About us Contact Privacy Terms