Antibodies, Proteins, Kits and Reagents for Life Science

314 937 ₸ Product size

100 µl 314 937 ₸

Order now and get it on Tuesday March 02, 2021

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EP2419Y] to IRF3
Suitable for: WB, IP, IHC-P, Flow Cyt
Knockout validated
Reacts with: Human

Product name

Anti-IRF3 antibody [EP2419Y]
See all IRF3 primary antibodies
Description

Rabbit monoclonal [EP2419Y] to IRF3
Host species

Rabbit

Tested Applications & Species

Application	Species
Flow Cyt	Human
IHC-P	Human
IP	Human
WB	Human

See all applications and species data

Immunogen

Synthetic peptide within Human IRF3 (C terminal). The exact sequence is proprietary.
Positive control
- WB: A549, MCF7, HeLa, HAP1, U-937, THP-1,Daudi and Jurkat cell lysates. IP: HeLa whole cell lysate. IHC: Human bladder carcinoma tissue. Flow cyt: HeLa cells.
General notes
Mouse, Rat: We have preliminary internal testing data to indicate this antibody may not react with these species. Please contact us for more information.
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.
We are constantly working hard to ensure we provide our customers with best in class antibodies. As a result of this work we are pleased to now offer this antibody in purified format. We are in the process of updating our datasheets. The purified format is designated 'PUR' on our product labels. If you have any questions regarding this update, please contact our Scientific Support team.

Form

Liquid
Storage instructions

Shipped at 4°C. Upon delivery aliquot and store at -20°C. Avoid freeze / thaw cycles.
Storage buffer

pH: 7.20
Preservative: 0.01% Sodium azide
Constituents: 0.05% BSA, 40% Glycerol (glycerin, glycerine), 59% PBS
Concentration information loading...
Purity

Protein A purified
Clonality

Monoclonal
Clone number

EP2419Y
Isotype

IgG
Research areas

Flow Cytometry - Anti-IRF3 antibody [EP2419Y] (ab76409)

Flow Cytometry analysis of HeLa (Human cervix adenocarcinoma epithelial cell) cells labeling IRF3 with Purified 76409 at 1:20 dilution (10 µg/ml) (Red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor® 488,ab150077) secondary antibody was used at 1:2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-IRF3 antibody [EP2419Y] (ab76409)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human bladder carcinoma tissue sections labeling IRF3 with Purified 76409 at 1:100 dilution (1.55 µg/ml). Heat mediated antigen retrieval using Bond™ Epitope Retrieval Solution 2 (pH 9.0) . Rabbit specific IHC polymer detection kit HRP/DAB (ab209101) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.
Western blot - Anti-IRF3 antibody [EP2419Y] (ab76409)

All lanes : Anti-IRF3 antibody [EP2419Y] (ab76409) at 1/1000 dilution (Purified)

Lane 1 : HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate
Lane 2 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate
Lane 3 : U-937 (Human histiocytic lymphoma monocyte) whole cell lysate
Lane 4 : THP-1 (Human monocytic leukemia monocyte) whole cell lysate
Lane 5 : Daudi (Human Burkitt's lymphoma lymphoblast) whole cell lysate

Lysates/proteins at 20 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 47 kDa

We are unsure how to define the extra bands.
Western blot - Anti-IRF3 antibody [EP2419Y] (ab76409)

All lanes : Anti-IRF3 antibody [EP2419Y] (ab76409) at 1/1000 dilution

Lane 1 : Jurkat cell lysate
Lane 2 : MCF7 cell lysate
Lane 3 : Wild-type HeLa cell lysate
Lane 4 : IRF3 knockout HeLa cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 47 kDa

Lanes 1 - 4: Merged signal (red and green). Green - ab76409 observed at 50 kDa. Red - loading control, ab8245 observed at 37 kDa.

ab76409 was shown to react with IRF3 in wild-type HeLa. Loss of signal was observed when knockout cell line ab255345 (knockout cell lysate ab263784) was used. Wild-type and IRF3 knockout samples were subjected to SDS-PAGE. ab76409 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.
Immunoprecipitation - Anti-IRF3 antibody [EP2419Y] (ab76409)

Purified ab76409 at 1/20 dilution (0.8µg) immunoprecipitating IRF3 in HeLa whole cell lysate.
Lane 1 (input): HeLa (Human cervix adenocarcinoma epithelial cell) whole cell lysate 10µg
Lane 2 (+): ab76409 + HeLa whole cell lysate.
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab76409 in HeLa 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: 47 kDa
Fresh lysate need to be used to avoid protein degradation.
Western blot - Anti-IRF3 antibody [EP2419Y] (ab76409)

All lanes : Anti-IRF3 antibody [EP2419Y] (ab76409) at 1/1000 dilution

Lane 1 : Wild-type A549 cell lysate
Lane 2 : IRF3 knockout A549 cell lysate

Lysates/proteins at 20 µg per lane.

Performed under reducing conditions.

Predicted band size: 47 kDa
Observed band size: 50 kDa
why is the actual band size different from the predicted?

Lanes 1 - 2: Merged signal (red and green). Green - ab76409 observed at 50 kDa. Red - loading control ab8245 (Mouse anti-GAPDH antibody [6C5]) observed at 37kDa.

ab76409 was shown to react with IRF3 in wild-type A549 cells in western blot with loss of signal observed in IRF3 knockout cell line ab267097 (IRF3 knockout cell lysate ab256953). Wild-type and IRF3 knockout A549 cell lysates were subjected to SDS-PAGE. Membranes were blocked in 3% milk in TBS-T (0.1% Tween^®) before incubation with ab76409 and ab8245 (Mouse anti-GAPDH antibody [6C5]) overnight at 4°C at a 1 in 1000 dilution and a 1 in 20000 dilution respectively. Blots were incubated 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 in 20000 dilution for 1 hour at room temperature before imaging.
Western blot - Anti-IRF3 antibody [EP2419Y] (ab76409)

All lanes : Anti-IRF3 antibody [EP2419Y] (ab76409) at 1/1000 dilution

Lane 1 : Wild-type HAP1 cell lysate
Lane 2 : IRF3 knockout HAP1 cell lysate
Lane 3 : HeLa cell lysate
Lane 4 : Jurkat cell lysate

Lysates/proteins at 20 µg per lane.

Predicted band size: 47 kDa

Lanes 1 - 4: Merged signal (red and green). Green - ab76409 observed at 50 kDa. Red - loading control, ab8245, observed at 37kDa.

ab76409 was shown to react with IRF3 in wild-type HAP1 cells alond with additional cross-reactive bands. No band was observed when IRF3 knockout samples were used. Wild-type and IRF3 knockout samples were subjected to SDS-PAGE. ab76409 and ab8245 (loading control to GAPDH) were both diluted 1/1000 and 1/10,000 respectively and incubated overnight at 4°C. 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 1/10,000 dilution for 1 hour at room temperature before imaging.

Please note: All products are "FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES"
For licensing inquiries, please contact partnerships@abcam.com