Anti-BAG2 antibody [EPR3567]

• IHC-P

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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-BAG2 antibody [EPR3567] (AB79406)

ab79406, at 1/100 dilution, staining BAG2 in human ovarian carcinoma by Immunohistochemistry using paraffin-embedded tissue.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

• IP

Lab

Immunoprecipitation - Anti-BAG2 antibody [EPR3567] (AB79406)

BAG2 was immunoprecipitated from 0.35 mg Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 μg with 79406 at 1/50 dilution (2μg). VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution.

Lane 1 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 μg

Lane 2 : ab79406 IP in Jurkat whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab79406 in Jurkat whole cell lysate

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

All lanes:

Immunoprecipitation - Anti-BAG2 antibody [EPR3567] (ab79406)

Predicted band size: 23 kDa

Observed band size: 24 kDa

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• WB

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Western blot - Anti-BAG2 antibody [EPR3567] (AB79406)

All lanes:

Western blot - Anti-BAG2 antibody [EPR3567] (ab79406) at 1/5000 dilution

Lane 1:

Jurkat cell lysate at 10 µg

Lane 2:

HeLa cell lysate at 10 µg

Lane 3:

HepG2 cell lysate at 10 µg

Secondary

All lanes:

Goat anti-rabbit HRP at 1/2000 dilution

Predicted band size: 23 kDa

Observed band size: 24 kDa

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• WB

Lab

Western blot - Anti-BAG2 antibody [EPR3567] (AB79406)

Lanes 1-3 : Merged signal (red and green). Green - ab79406 observed at 25 kDa. Red - loading control ab8245 observed at 36 kDa.

ab79406 Anti-BAG2 antibody [EPR3567] was shown to specifically react with BAG2 in wild-type HeLa cells. Loss of signal was observed when knockout cell line ab265907 (knockout cell lysate ab257369) was used. Wild-type and BAG2 knockout samples were subjected to SDS-PAGE. ab79406 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-BAG2 antibody [EPR3567] (ab79406) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

BAG2 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human BAG2 knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-bag2-knockout-hela-cell-line-ab265907'>ab265907</a>)

Lane 3:

HepG2 cell lysate at 20 µg

Secondary

All lanes:

Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-irdye-800cw-preadsorbed-ab216773'>ab216773</a>) at 1/10000 dilution

Predicted band size: 23 kDa

Observed band size: 25 kDa

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• WB

CiteAb

Western blot - Anti-BAG2 antibody [EPR3567] (AB79406)

BAG2 western blot using anti-BAG2 antibody [EPR3567] ab79406. Publication image and figure legend from Wang, C., Scott, S. M., et al., 2019, Nat Commun, PubMed 31699992.

ab79406 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 ab79406 please see the product overview.

Hyperacetylation of NPC1 and proteostasis impact by SAHA. a SAHA leads hyperacetylation of both WT and I1061T-NPC1. Cell lysate of WT or I1061T/I1061T patient fibroblast was immunoprecipitated by acetylated-lysine (AcK) antibody and then recognized by NPC1 antibody through western blot. b SAHA impacts the expression of proteostasis components. Immunoblot analysis of SAHA-treated (10 μM for 48 h) I1061T/I1061T homozygous fibroblast (left panel) and quantification of total Hsf1, Hsf1 phosphorylated (Hsf1-P), BAG1, BAG2, BAG3, and HDAC7 (right panel) are shown. GAPDH was used as Western blot loading control. Data is presented as fold change to DMSO treatment (mean ± s.d., n = 3). P-values are indicated using student's two tailed t-test (*p < 0.05, **p<0.01, ***p<0.001)

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