Anti-Bag1 antibody [Y166]

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Bag1 antibody [Y166] (AB32109)

Immunofluorescent staining of HeLa cells using ab32109 at 1/100 dilution.

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bag1 antibody [Y166] (AB32109)

Immunohistochemical analysis of paraffin-embedded human prostate carcinoma using ab32109 at 1/100 dilution.

Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

• WB

Lab

Western blot - Anti-Bag1 antibody [Y166] (AB32109)

Western blot : Anti-Bag1 antibody [Y166] ab32109 staining at 1/500 dilution, shown in green; Mouse anti alpha Tubulin ab7291 loading control staining at 1/20000 dilution, shown in magenta. A band was observed at 32 kDa in Wild-type A549 cell lysates with no signal observed at this size in BAG1 knockout A549 cell line. To generate this image, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3pc 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 800CW and Goat anti-Mouse 680RD at 1/20,000 dilution.

All lanes:

Western blot - Anti-Bag1 antibody [Y166] (ab32109) at 1/500 dilution

Lane 1:

Wild-type A549 at 20 µg

Lane 2:

BAG1 knockout A549 at 20 µg

Lane 3:

THP-1 at 20 µg

Lane 4:

HT1080 at 20 µg

Secondary

All lanes:

Goat anti-Rabbit 800CW & Goat anti-Mouse 680RD at 1/20000 dilution

Predicted band size: 39 kDa

Observed band size: 32 kDa,51 kDa

true

Exposure time: 8s

• WB

Unknown

Western blot - Anti-Bag1 antibody [Y166] (AB32109)

All lanes:

Western blot - Anti-Bag1 antibody [Y166] (ab32109) at 1/500 dilution

All lanes:

HeLa cell lysate

Predicted band size: 39 kDa

Observed band size: 33 kDa,46 kDa,50 kDa

false

• WB

CiteAb

Western blot - Anti-Bag1 antibody [Y166] (AB32109)

Bag1 western blot using anti-Bag1 antibody [Y166] ab32109. Publication image and figure legend from Wood, R. J., Ormsby, A. R., et al., 2018, Nat Commun, PubMed 29348634.

ab32109 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 ab32109 please see the product overview.

Probing changes in proteostasis through selective activation of different chaperone systems. a Investigation of the effect of induction of the heat shock response by HSF1 overexpression or selective overexpression of HSP90. Shown on left are western blots of the HEK293T cells matched for total protein via a BCA assay. These cells were co-transfected with HSF1 (or myc-tagged HSP90 or control of (non-fluorescent) Y66L Emerald fluorescent protein) and the L89G barnase biosensor (which also contains a myc tag). The right graphs show the Lower-slope and A50% analyses of these treatments and one or three of the biosensor variants as indicated vs. the wild-type* variant. Bars indicate means ± SEM. The right panels were analysed via a two-way ANOVA subjected to a Dunnett's post-hoc test, results coded as ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, ns = > 0.05. b Same paradigm as panel a. In this case, BAG1 was cotransfected with the barnase biosensor, HSP40 (DNAJB1) and HSP70 (HSPA1A). The right panels show the proportional dosage of each construct (by mass of DNA) in the transfection. Bars indicate means ± SEM. The right panel Lower-slope graph was analysed via a two-way ANOVA subjected to Dunnett's post hoc test and the A50% graph was analysed via a one-way ANOVA subjected to a Tukey's post-hoc test. Results coded as ****p < 0.0001, ***p < 0.001, **p < 0.01, *p < 0.05, ns = > 0.05

false