Anti-Beclin 1 antibody [EPR20473]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Immunohistochemical analysis of paraffin-embedded human breast cancer tissue labeling Beclin 1 with ab210498 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Granularly cytoplasmic staining on tumor cells of human breast cancer is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

We don't recommend this antibody for mouse and rat in IHC. In our hands most mouse and rat tissues showed nuclear staining.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Immunohistochemical analysis of paraffin-embedded human kidney tissue labeling Beclin 1 with ab210498 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

Granularly cytoplasmic staining on human kidney is observed.

Counter stained with Hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) Ready to use.

We don't recommend this antibody for mouse and rat in IHC. In our hands most mouse and rat tissues showed nuclear staining.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

• IHC-P

Lab

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Immunohistochemistry analysis of paraffin embedded sections of human hepatocellular carcinoma tissue staining Beclin 1 with ab210498 at 1/100 dilution and ready to use secondary Goat Anti-Rabbit IgG H&L (HRP polymer) (ab214880). Counterstained with hematoxylin. Heat mediated antigen retrieval was performed using ab93684 (Tris/EDTA buffer, pH 9.0). Positive staining on human hepatocellular carcinoma. The section was incubated with ab210498 at 4°C overnight.

• IP

Collaborator

Immunoprecipitation - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Immunoprecipitation of BECN1 in HeLa cells. Lysates were prepared and immunoprecipitation was performed using 2 μg of ab210498 pre-coupled to Protein A beads. Samples were then washed and processed for western blot.

This data was provided by YCharOS Inc., an open science company with the mission of characterizing commercially available antibody reagents for all human proteins. Abcam and YCharOS are working together to help address the reproducibility crisis by enabling the life science community to better evaluate commercially available antibodies.

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• IP

Supplier Data

Immunoprecipitation - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Beclin 1 was immunoprecipitated from 0.35 mg of HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate with ab210498 at 1/30 dilution.

Western blot was performed from the immunoprecipitate using ab210498 at 1/1000 dilution.

VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

Lane 1 : HepG2 whole cell lysate, 10 μg (Input).

Lane 2 : ab210498 IP in HepG2 whole cell lysate.

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab210498 in HepG2 whole cell lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.

Exposure time : 30 seconds.

All lanes:

Immunoprecipitation - Anti-Beclin 1 antibody [EPR20473] (ab210498)

Predicted band size: 52 kDa

Observed band size: 52 kDa

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• WB

Supplier Data

Western blot - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Beclin 1 antibody [EPR20473] (ab210498) at 1/1000 dilution

Lane 1:

Human fetal brain lysate at 10 µg

Lane 2:

Human colon lysate at 10 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/4000 dilution

Predicted band size: 52 kDa

Observed band size: 52 kDa

false

Exposure time: 3min

• WB

Lab

Western blot - Anti-Beclin 1 antibody [EPR20473] (AB210498)

False colour image of Western blot : Anti-Beclin 1 antibody [EPR20473] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab210498 was shown to bind specifically to Beclin 1. A band was observed at 52 kDa in wild-type Jurkat cell lysates with no signal observed at this size in BECN1 CRISPR-Cas9 edited cell line ab283778 (BECN1 CRISPR-Cas9 edited cell lysate ab283792). The band observed in the CRISPR-Cas9 edited lysate lane above 52 kDa is likely to represent Beclin 1 with an insertion. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and BECN1 CRISPR-Cas9 edited Jurkat cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) before incubation with primary antibodies overnight at 4 °C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-Beclin 1 antibody [EPR20473] (ab210498) at 1/1000 dilution

Lane 1:

Wild-type Jurkat cell lysate at 20 µg

Lane 2:

BECN1 CRISPR-Cas9 edited Jurkat cell lysate at 20 µg

Lane 3:

HeLa cell lysate at 20 µg

Lane 4:

Raji cell lysate at 20 µg

Secondary

Lanes 1 - 4:

Goat anti-Rabbit IgG H&L 800CW at 1/20000 dilution

Lanes 1 - 4:

Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution

Observed band size: 52 kDa

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• WB

Supplier Data

Western blot - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Beclin 1 antibody [EPR20473] (ab210498) at 1/1000 dilution

Lane 1:

HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate at 20 µg

Lane 2:

HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate at 20 µg

Lane 3:

HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate at 20 µg

Lane 4:

T-47D (Human ductal breast epithelial tumor cell line) whole cell lysate at 20 µg

Lane 5:

Raji (Human Burkitt's lymphoma cell line) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 52 kDa

Observed band size: 52 kDa

false

Exposure time: 15s

• WB

Supplier Data

Western blot - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Beclin 1 antibody [EPR20473] (ab210498) at 1/1000 dilution

Lane 1:

RAW 264.7 (Mouse macrophage cell line transformed with Abelson murine leukemia virus) whole cell lysate at 10 µg

Lane 2:

NIH/3T3 (Mouse embryonic fibroblast cell line) whole cell lysate at 10 µg

Lane 3:

A20 (Mouse reticulum sarcoma cell line) whole cell lysate at 10 µg

Lane 4:

C2C12 (Mouse myoblast cell line) whole cell lysate

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 52 kDa

Observed band size: 52 kDa

false

Exposure time: 1min

• WB

Supplier Data

Western blot - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Western blot - Anti-Beclin 1 antibody [EPR20473] (ab210498) at 1/1000 dilution

Lane 1:

Mouse brain lysate at 10 µg

Lane 2:

Mouse spleen lysate at 10 µg

Lane 3:

Rat brain lysate at 10 µg

Lane 4:

Rat kidney lysate at 10 µg

Lane 5:

Rat spleen lysate at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 52 kDa

Observed band size: 52 kDa

false

Exposure time: 3min

• WB

CiteAb

Western blot - Anti-Beclin 1 antibody [EPR20473] (AB210498)

Beclin 1 western blot using anti-Beclin 1 antibody [EPR20473] ab210498. Publication image and figure legend from Chow, J. P. H., Cai, Y., et al., 2020, PLoS One, PubMed 32353864.

ab210498 was used in this publication in western blot. This may not be the same as the application(s) guaranteed by Abcam. For a full list of applications guaranteed by Abcam for ab210498 please see the product overview.

Expression of ASS1 in various cell lines and the effect of NEI-01 on their cell viability.(A) Expression of ASS1, ASL and OTC in SMMC7721, MIA PaCa-2 and PANC-1. Unperturbed asynchronously growing cells were harvested by trypsinization. After cell lysis, total cell lysates were subject to immunoblot with anti-ASS1, anti-ASL, anti-OTC or anti-ACTIN antibodies. (B) Inhibition of cell growth of MIA PaCa-2 and PANC-1 cells but not SMMC7721 cells by NEI-01 treatment. SMMC7721 cells, MIA PaCa-2 cells or PANC-1 were seeded in 96 well-plate with ~3000 cells/well density 24 hr prior to NEI-01 treatments. After 72 hr, cells were subject to MTT assay. The relative cell viability in % vs the NEI-01 concentration was shown as mean ± SD of triplicate wells. (C) Activation of autophagic pathway upon NEI-01 treatment. MIA PaCa-2 cells were treated with indicated amount of NEI-01 for 3 or 7 days. The cells were then harvested and subject to immunoblot using designated antibodies.

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