Antibodies, Proteins, Kits and Reagents for Life Science

355 142 ₸ Product size

100 µl 355 142 ₸

Order now and get it on Tuesday March 02, 2021

Anti-ATF-4 antibody [EPR18111] (ab184909)

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR18111] to ATF-4
Suitable for: WB, IHC-P, ICC/IF, Flow Cyt, IP
Reacts with: Human

Product name

Anti-ATF-4 antibody [EPR18111]
See all ATF-4 primary antibodies
Description

Rabbit monoclonal [EPR18111] to ATF-4
Host species

Rabbit

Tested Applications & Species

Application	Species
Flow Cyt	Human
ICC/IF	Human
IHC-P	Human
IP	Human
WB	Human

See all applications and species data

Immunogen

Recombinant fragment. This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: Human fetal liver and fetal kidney lysates; A431, HeLa, whole cell lysates; untransfected HEK-293 whole cell lysate (control) lysate; HEK-293 whole cell lysate transfected with ATF-4 with GFP-tag lysate; HepG2 treated with 5 µM MG-132 (ab141003) and 3 µg/ml tunicamycin (ab120296) for 6 hours whole cell lysate. IHC-P: Human bladder cancer and stomach cancer tissues. ICC/IF: HeLa and A431 cells. Flow Cyt: HeLa cells. IP: HeLa whole cell lysate.
General notes
This product is a recombinant monoclonal antibody, which offers several advantages including:
- - High batch-to-batch consistency and reproducibility
- - Improved sensitivity and specificity
- - Long-term security of supply
- - Animal-free production
For more information see here.
Our RabMAb^® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb^® patents.

Form

Liquid
Storage instructions

Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle.
Storage buffer

pH: 7.2
Preservative: 0.01% Sodium azide
Constituents: 59% PBS, 40% Glycerol, 0.05% BSA
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EPR18111
Isotype

IgG
Research areas

Western blot - Anti-ATF-4 antibody [EPR18111] (ab184909)

All lanes : Anti-ATF-4 antibody [EPR18111] (ab184909) at 1/1000 dilution

Lane 1 : Human fetal liver lysate
Lane 2 : A431 (Human epidermoid carcinoma cell line) whole cell lysate
Lane 3 : HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate
Lane 4 : Human fetal kidney lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG Peroxidase Conjugate, specific to the non-reduced form of IgG at 1/100000 dilution

Predicted band size: 38 kDa
Observed band size: 50 kDa
why is the actual band size different from the predicted?

Blocking/Dilution buffer: 5% NFDM/TBST.

Exposure time: Lane 1: 3 minutes; Lane 2 and 3: 30 seconds; Lane 4: 3 minutes.
Immunocytochemistry/ Immunofluorescence - Anti-ATF-4 antibody [EPR18111] (ab184909)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling ATF-4 with ab184909 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on HeLa cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows:-

-ve control 1: ab184909 at 1/1000 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2: ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATF-4 antibody [EPR18111] (ab184909) This image was courtesy of an annoymous AbReview

Formalin-fixed paraffin-embedded human colon cancer tissue stained for ATF-4 using ab184909 at 1/80 dilution (45 minute incubation) in immunohistochemical analysis. A polyclonal Goat Horse Radish Peroxidase antibody was used as the secondary.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATF-4 antibody [EPR18111] (ab184909)

Immunohistochemical analysis of paraffin-embedded human stomach cancer tissue labeling ATF-4 with ab184909 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on human stomach 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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Western blot - Anti-ATF-4 antibody [EPR18111] (ab184909)

All lanes : Anti-ATF-4 antibody [EPR18111] (ab184909) at 1/5000 dilution

Lane 1 : Untransfected HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate (control)
Lane 2 : HEK-293 (Human epithelial cell line from embryonic kidney) whole cell lysate transfected with ATT-4 with GFP-tag lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 38 kDa
Observed band size: 50 kDa why is the actual band size different from the predicted?

Exposure time: 3 minutes

Blocking/Dilution buffer: 5% NFDM/TBST.
Western blot - Anti-ATF-4 antibody [EPR18111] (ab184909)

All lanes : Anti-ATF-4 antibody [EPR18111] (ab184909) at 1/1000 dilution

Lane 1 : Untreated HepG2 (Human liver hepatocellular carcinoma cell line) whole cell lysate (control)
Lane 2 : HepG2 (Human liver hepatocellular carcinoma cell line) treated with 5 µM MG-132 (ab141003) and 3 µg/ml tunicamycin (ab120296) for 6 hours whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

Predicted band size: 38 kDa
Observed band size: 50 kDa why is the actual band size different from the predicted?

Exposure time: 15 seconds

Blocking/Dilution buffer: 5% NFDM/TBST.

The molecular weight observed is consistent with what has been described in the literature (PMID: 22095285).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ATF-4 antibody [EPR18111] (ab184909)

Immunohistochemical analysis of paraffin-embedded human bladder cancer tissue labeling ATF-4 with ab184909 at 1/50 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/500 dilution.

Nucleus staining on human bladder 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) (ab97051) at 1/500 dilution.

Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
Immunocytochemistry/ Immunofluorescence - Anti-ATF-4 antibody [EPR18111] (ab184909)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized A431 (Human epidermoid carcinoma cell line) cells labeling ATF-4 with ab184909 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

Confocal image showing nuclear staining on A431 cell line.

The nuclear counterstain is DAPI (blue).

Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Loading Control (ab7291) at 1/1000 dilution and Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) preadsorbed (ab150120) at 1/1000 dilution (red).

The negative controls are as follows:-

-ve control 1: ab184909 at 1/1000 dilution followed by ab150120 at 1/1000 dilution.

-ve control 2: ab7291 at 1/1000 dilution followed by ab150077 at 1/1000 dilution.
Flow Cytometry - Anti-ATF-4 antibody [EPR18111] (ab184909)

Flow cytometric analysis of 4% paraformaldehyde-fixed HeLa (Human epithelial cell line from cervix adenocarcinoma) cells labeling ATF-4 with ab184909 at 1/80 dilution (red) compared with a Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (black) and an unlabelled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat Anti-Rabbit IgG (Alexa Fluor® 488) at 1/500 dilution was used as the secondary antibody.
Immunoprecipitation - Anti-ATF-4 antibody [EPR18111] (ab184909)

ATF-4 was immunoprecipitated from 1mg of HeLa (Human epithelial cell line from cervix adenocarcinoma) whole cell lysate with ab184909 at 1/50 dilution.

Western blot was performed from the immunoprecipitate using ab184909 at 1/1000 dilution.

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

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

Lane 2: ab184909 IP in HeLa whole cell lysate.

Lane 3: Rabbit IgG,monoclonal [EPR25A] - Isotype Control (ab172730) instead of ab184909 in HeLa whole cell lysate

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

Exposure time: 3 minutes.
Anti-ATF-4 antibody [EPR18111] (ab184909)

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