Anti-Caspase-7 antibody [E22]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Caspase-7 antibody [E22] (AB32522)

Immunohistochemical analysis of paraffin embedded human skin cancer tissue using ab32522 at 1 : 50 dilution.

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

• Flow Cyt (Intra)

Unknown

Flow Cytometry (Intracellular) - Anti-Caspase-7 antibody [E22] (AB32522)

Intracellular Flow Cytometry analysis of HeLa (human cervix adenocarcinoma) cells labeling Caspase-7 (red) with ab32522 at a 1/250 dilution. Cells were fixed with 4% paraformaldehyde and permeabilized with 90% methanol. A goat anti-rabbit IgG (Alexa Fluorr^® 488) (ab150077) was used as the secondary antibody at a 1/2000 dilution. Black - Rabbit monoclonal IgG (ab172730). Blue (unlabeled control) - Cells without incubation with the primary and secondary antibodies.

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-Caspase-7 antibody [E22] (AB32522)

Immunofluorescent staining of HeLa cells using ab32522 at 1 : 100 dilution.

• IP

Lab

Immunoprecipitation - Anti-Caspase-7 antibody [E22] (AB32522)

Purified ab32522 at 1/20 dilution (1μg) immunoprecipitating Caspase-7 in Jurkat whole cell lysate.
Lane 1 (input) : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10μg
Lane 2 (+) : ab32522 + Jurkat whole cell lysate.
Lane 3 (-) : Rabbit monoclonal IgG (ab172730) instead of ab32522 in Jurkat whole cell lysate.
VeriBlot for IP Detection Reagent (HRP) (ab131366) (1/1000 dilution) was used for Western blotting.
Blocking Buffer and concentration : 5% NFDM/TBST.
Diluting buffer and concentration : 5% NFDM/TBST.
Observed band size : 34 kDa

All lanes:

Immunoprecipitation - Anti-Caspase-7 antibody [E22] (ab32522)

Predicted band size: 34 kDa

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• WB

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Western blot - Anti-Caspase-7 antibody [E22] (AB32522)

Lanes 1- 2 : Merged signal (red and green). Green - ab32522 observed at 38 kDa. Red - Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) observed at 50 kDa.

ab32522 was shown to react with pro Caspase-7 in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab265777 (knockout cell lysate ab257380) was used. Wild-type HeLa and CASP7 knockout HeLa cell lysates were subjected to SDS-PAGE. ab32522 and Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) overnight at 4°C at a 1 in 1000 dilution and a 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-Caspase-7 antibody [E22] (ab32522) at 1/1000 dilution

Lane 1:

Wild-type HeLa cell lysate at 20 µg

Lane 2:

CASP7 knockout HeLa cell lysate at 20 µg

Lane 2:

Western blot - Human CASP7 (Caspase-7) knockout HeLa cell line (<a href='/en-us/products/cell-lines/human-casp7-caspase-7-knockout-hela-cell-line-ab265777'>ab265777</a>)

Predicted band size: 34 kDa

Observed band size: 38 kDa

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• WB

Lab

Western blot - Anti-Caspase-7 antibody [E22] (AB32522)

Lane 1 : Wild type HAP1 whole cell lysate (20 μg)
Lane 2 : Wild type HAP1 + Staurosporine ab120056 whole cell lysate (20 μg)
Lane 3 : CASP7 knockout HAP1 whole cell lysate (20 μg)
Lane 4 : CASP7 + Staurosporine knockout HAP1 whole cell lysate (20 μg)
Lane 5 : HeLa whole cell lysate (20 μg)
Lane 6 : HeLa + Staurosporine whole cell lysate (20 μg)

Lanes 1 - 6 : Merged signal (red and green). Green - ab32522 observed at 38 kDa. Red - loading control, ab18058, observed at 130 kDa.

ab32522 was shown to specifically react with HAP1 + Staurosporine when HAP1 + Staurosporine knockout samples were used. Wild-type and HAP1 + Staurosporine knockout samples were subjected to SDS-PAGE. ab32522 and ab18058 (Mouse anti Vinculin loading control) were incubated overnight at 4°C at 1000 dilution and 1/10000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye^® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye^® 680RD) preabsorbed ab216776 secondary antibodies at 1/20000 dilution for 1 hour at room temperature before imaging.

All lanes:

Western blot - Anti-Caspase-7 antibody [E22] (ab32522)

Predicted band size: 34 kDa

false

• WB

Unknown

Western blot - Anti-Caspase-7 antibody [E22] (AB32522)

All lanes:

Western blot - Anti-Caspase-7 antibody [E22] (ab32522) at 1/1000 dilution

All lanes:

Jurkat cell lysate

Predicted band size: 34 kDa

Observed band size: 34 kDa

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