重组Anti-FRA1抗体[EPR23767-177]

• Flow Cyt (Intra)

Supplier Data

Flow Cytometry (Intracellular) - Anti-FRA1 antibody [EPR23767-177] (AB252421)

Flow cytometric analysis of 4% paraformaldehyde-fixed, 90% methanol-permeabilized HeLa (human cervix adenocarcinoma epithelial cell) starved for 24 hours then treated with 100 ng/ml TPA for 4 hours (Red) /Untreated control (Green) cells labelling FRA1 with ab252421 at 1/500 dilution (0.1ug) compared with a Rabbit monoclonal IgG (ab172730) (Black) isotype control and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (Blue).

A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) at 1/2000 dilution was used as the secondary antibody.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-FRA1 antibody [EPR23767-177] (AB252421)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa cells labelling FRA1 with ab252421 at 1/500 (0.918 ug/ml) dilution, followed by ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) antibody at 1/1000 dilution (Green). The expression of FRA1 proteins is induced by TPA treatment (PMID : 13679379). Confocal image showing nuclear staining in HeLa cell line after starvation 24 hours, then treatment with Phorbol-12-myristate-13-acetate for 4h is observed. ab195889 Anti-alpha Tubulin mouse monoclonal antibody - Microtubule Marker (Alexa Fluor^® 594) was used to counterstain tubulin at 1/200 dilution (Red). The Nuclear counterstain was DAPI (Blue).

Secondary antibody only control : Secondary antibody is ab150077 Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) at 1/1000 dilution.

• IP

Supplier Data

Immunoprecipitation - Anti-FRA1 antibody [EPR23767-177] (AB252421)

FRA1 was immunoprecipitated from 0.35 mg HeLa (human cervix adenocarcinoma epithelial cell) staved for 24 hours then treated with 100 ng/mL TPA for 4 hours, whole cell lysate with ab252421 at 1/30 dilution (2ug in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab252421 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366) was used at 1/5000 dilution.

Lane 1 : HeLa (human cervix adenocarcinoma epithelial cell) staved for 24 hours then treated with 100 ng/mL TPA for 4 hours, whole cell lysate 10 ug

Lane 2 : ab252421 IP in HeLa staved for 24 hours then treated with 100 ng/mL TPA for 4 hours whole cell lysate

Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab252421 in HeLa staved for 24 hours then treated with 100 ng/mL TPA for 4 hours whole cell lysate

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

Exposure time : 15 seconds.

All lanes:

Immunoprecipitation - Anti-FRA1 antibody [EPR23767-177] (ab252421)

Predicted band size: 29 kDa

Observed band size: 40 kDa

false

• WB

Lab

Western blot - Anti-FRA1 antibody [EPR23767-177] (AB252421)

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

The molecular weight observed is consistent with what has been described in the literature (PMID : 13679379).

The expression of FRA1 proteins is induced by TPA treatment (PMID : 13679379).

Exposure time : 3 minutes.

All lanes:

Western blot - Anti-FRA1 antibody [EPR23767-177] (ab252421) at 1/1000 dilution

Lane 1:

HeLa (human cervix adenocarcinoma epithelial cell) staved for 24 hours, whole cell lysate at 20 µg

Lane 2:

HeLa staved for 24 hours then treated with 100 ng/mL TPA (<a href='/products/biochemicals/phorbol-12-myristate-13-acetate-pma-pkc-activator-ab120297'>ab120297</a>) for 4 hours, whole cell lysate at 20 µg

Lane 3:

NIH/3T3 (mouse embryonic fibroblast) staved for 2 hours, whole cell lysate at 20 µg

Lane 4:

NIH/3T3 staved for 2 hours then treated with 100 ng/mL TPA (<a href='/products/biochemicals/phorbol-12-myristate-13-acetate-pma-pkc-activator-ab120297'>ab120297</a>) for 4 hours, whole cell lysate at 20 µg

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/50000 dilution

Predicted band size: 29 kDa

Observed band size: 40 kDa

false

• WB

Lab

Western blot - Anti-FRA1 antibody [EPR23767-177] (AB252421)

Blocking and diluting buffer and concentration : 5% NFDM/TBST

Exposure time : 3 minutes.

All lanes:

Western blot - Anti-FRA1 antibody [EPR23767-177] (ab252421) at 1/1000 dilution

All lanes:

MDA-MB-231 (human breast adenocarcinoma epithelial cell) whole cell lysate at 40 µg

Secondary

All lanes:

VeriBlot for IP secondary antibody(HRP)(<a href='/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/1000 dilution

Predicted band size: 29 kDa

Observed band size: 40 kDa

false