Antibodies, Proteins, Kits and Reagents for Life Science

304 886 ₸ Product size

100 µl 304 886 ₸

Order now and get it on Tuesday March 09, 2021

Key features and details

Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
Rabbit monoclonal [EPR5249] to KAP1
Suitable for: IP, Flow Cyt, WB, IHC-P, ICC/IF
Knockout validated
Reacts with: Mouse, Human

Product name

Anti-KAP1 antibody [EPR5249]
See all KAP1 primary antibodies
Description

Rabbit monoclonal [EPR5249] to KAP1
Host species

Rabbit
Specificity

Although the immunogen is from a phosphor-peptide, this antibody detects phospho and non-phospho KAP1. Based on a peptide blocking experiment it has been found that the signal generated after non-phospho peptide blocking became much weaker, thus indicating that ab109545 shows cross-reactivity with the non-phospho KAP1 at high level.

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

This information is proprietary to Abcam and/or its suppliers.
Positive control
- WB: A431 and MCF7 cell lysates; IHC-P: Human colon and kidney tissue; ICC/IF: Hela cells; IP: A431 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: 7.20
Preservative: 0.05% Sodium azide
Constituents: 0.1% BSA, 40% Glycerol (glycerin, glycerine), 9.85% Tris glycine, 50% Tissue culture supernatant
Concentration information loading...
Purity

Tissue culture supernatant
Clonality

Monoclonal
Clone number

EPR5249
Isotype

IgG
Research areas
- Cancer
- Cell Death
- Apoptosis
- Other
- Epigenetics and Nuclear Signaling
- Bromodomains

Western blot - Anti-KAP1 antibody [EPR5249] (ab109545)

Lane 1: Wild type HAP1 whole cell lysate (20 µg)
Lane 2: HAP1 + DMSO whole cell lysate (20 µg)
Lane 3: HAP1 + Blaomycin whole cell lysate (20 µg)
Lane 4: TRIM28 knockout HAP1 whole cell lysate (20 µg)
Lane 5: TRIM28 knockout HAP1 + DMSO whole cell lysate (20 µg)
Lane 6: TRIM28 knockout HAP1 + Blaomycin whole cell lysate (20 µg)
Lane 7: HeLa + DMSO whole cell lysate (20 µg)
Lane 8: HeLa + Blaomycin whole cell lysate (20 µg)

Lanes 1 - 8: Merged signal (red and green). Green - ab109545 observed at 110 kDa. Red - loading control, ab8245, observed at 37 kDa.

ab109545 was shown to specifically react with KAP1 in wild type cells as signal was lost in KAP1 knockout cells. Wild-type and KAP1 knockout samples were subjected to SDS-PAGE. ab109545 and ab8245 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 10000 dilution and 1/20000 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.
Immunocytochemistry/ Immunofluorescence - Anti-KAP1 antibody [EPR5249] (ab109545)

ab109545, at a 1/100 dilution, staining KAP1 in HeLa cells
Immunoprecipitation - Anti-KAP1 antibody [EPR5249] (ab109545)

KAP1 was immunoprecipitated from 0.35 mg A431 (Human epidermoid carcinoma epithelial cell) cell lysate 10 µg with ab109545 at 1/30 dilution (2µg in 0.35mg lysates). Western blot was performed on the immunoprecipitate using ab109545 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP)(ab131366) was used at 1/5000 dilution.
Lane 1: A431 (Human epidermoid carcinoma epithelial cell) cell lysate 10 µg

Lane 2: ab109545 IP in A431 cell lysate

Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab109545 in A431 cell lysate

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

Exposure time: 32 seconds
Western blot - Anti-KAP1 antibody [EPR5249] (ab109545)

All lanes : Anti-KAP1 antibody [EPR5249] (ab109545) at 1/10000 dilution

Lane 1 : A431 (Human epidermoid carcinoma epithelial cell) whole cell lysates
Lane 2 : MCF7 (Human breast adenocarcinoma epithelial cell) whole cell lysates

Lysates/proteins at 15 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/20000 dilution (Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated)

Developed using the ECL technique.

Predicted band size: 89 kDa
Observed band size: 110 kDa
why is the actual band size different from the predicted?

Exposure time: 10 seconds

Blocking/diluting buffer and concentration: 5% NFDM/TBST
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KAP1 antibody [EPR5249] (ab109545)

ab109545, at a 1/100 dilution, staining KAP1 in formalin-fixed, paraffin-embedded Human colon tissue.

Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-KAP1 antibody [EPR5249] (ab109545)

ab109545, at a 1/100 dilution, staining KAP1 in formalin-fixed, paraffin-embedded Human kidney tissue.

Perform heat mediated antigen retrieval via the pressure cooker method before commencing with IHC staining protocol.
Flow Cytometry - Anti-KAP1 antibody [EPR5249] (ab109545)

Overlay histogram showing HeLa cells stained with ab109545 (red line). The cells were fixed with 80% methanol (5 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab190545, 1/100 dilution) for 30 min at 22ºC. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22ºC. Isotype control antibody (black line) was rabbit IgG (monoclonal) (1µg/1x10⁶ cells) used under the same conditions. Acquisition of >5,000 events was performed.
Anti-KAP1 antibody [EPR5249] (ab109545)

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