Anti-Beclin 1 antibody [2A4] (ab114071)

Protocol tips

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	Incubate the membrane with appropriate dilutions of primary antibody in blocking buffer and incubate overnight at 4°C;

Protocol tips
Incubate the membrane with appropriate dilutions of primary antibody in blocking buffer and incubate overnight at 4°C;

Publication protocol

Whole cell lysates were further processed by SDS-Page followed by Western blotting, as previously described [15]. Immunodetection with primary antibodies against human Survivin (AF886, R&D Systems, Wiesbaden-Nordenstadt, Germany), Bcl-2 (610539, BD Transduction Laboratories) Beclin1 (ab114071), Map1lc3b (ab51520) and Sqstm1 (ab96706) (AbCam), DRAM1 (PRS4035 Sigma-Aldrich Saint Louis MO USA), ATF4 (ab50546, AbCam) and β-actin (A5441) (Sigma-Aldrich) was performed. Bound secondary HRP-conjugated anti-mouse (A9917) and anti-rabbit (A0545) antibodies (Sigma-Aldrich) were detected by incubating the immunoblots with Super Signal West Pico Chemiluminescent Substrate (#34077, Pierce, Thermo Fisher Scientific, Darmstadt Germany). The luminescent reactivity was then measured using Fusion image capture and further quantified with Bio1D analysis system (PEQLAB Biotechnologie GmbH). The blotted nitrocellulose membranes (Amersham Protran Premium 0.2 μm NC Cat. N. 10600009 Ge Healthcare Life Science, Freiburg Germany) were up to four times stripped with Stripping Buffer (Restore Plus Western Blot Stripping Buffer Cat. N. 46430 Thermo Scientific) as suggested by the company and reprobed in order to detect other proteins. At last, anti-β-actin was used to control equal loading and protein quality.

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Manufacturer protocol

Download the product protocol from Abcam for Anti-Beclin 1 antibody [2A4] (ab114071) below.

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