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Primary Antibodies  >  Sample Size Primary Antibodies

NF-kB p65 Antibody

Mouse Monoclonal 27F9.G4 IgG2a kappa


25 µL


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Rockland Monoclonal anti NFKB p65 (Rel A) antibody was used to detect p65 by immunofluorescence at a dilution of 1:5000.   Hela cells were grown to sub-confluent on 18 mm2 glass coverslips #1.5.  Cells were either unstimulated (A), or stimulated (B) with 50 ng/ml of TNF alpha for 30 min prior fixation.  Cells were then fixed in methanol and blocked with 10% normal goat serum (NGS), in PBS, and TritonX 0.2% (Tx) and incubated for 1 hr at RT with primary ab, counterstained with DAPI and washed in PBS/NGS/Tx.  Cells were incubated for 1 hr at RT with Atto 425 conjugated anti mouse secondary antibody for STED CW imaging.  Data was collected on a STED-CW TCS-SP5 Confocal system equipped with a DFC 350FX camera allowing sequential acquisition in widefiled, confocal and STED CW imaging on the same system.
Rockland anti NFKB p65 (Rel A) monoclonal antibody 200-301-065 was used to detect ~65 kD band (red arrow) in HeLa whole cell lysate.  Lysate was run on 4-20% gradient gel transferred under standard conditions and blocked in 1% BSA-TTBS 30 min RT.  Blot was probed with monoclonal anti p65 1:1000 in 1% BSA-TBS-T o/n 4°C and detected with HRP conjugated Rb-anti Mouse antibody 1:40,000 in MB-070 30 min RT.
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Synonyms: NFKB, nfkb, NF-kB, NF-kappaB, NFkappaB, Nuclear factor NF-kappa-B p65 subunit

NF-kB p65 Antibody Properties

Anti-NFKB p65 (Rel A) (MOUSE) Monoclonal Antibody - 200-301-065S
Target Species
Known Cross Reactivity
Monoclonal 27F9.G4 IgG2a kappa
ELISA : 1:50,000-1:100,000
IF Microscopy : 1:5000
Western Blot : 1:1000 - 1:5000
Immunohistochemistry: 1:200-1:600
Other Dilution: User Optimized
Physical State
Liquid (sterile filtered)
Shipping condition
Dry Ice
1.0 mg/mL by UV absorbance at 280 nm
0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2
0.01% (w/v) Sodium Azide
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NF-kB p65 Antibody Description

NFkappaB was originally identified as a factor that binds to the immunoglobulin kappa light chain enhancer in B cells. It was subsequently found in non-B cells in an inactive cytoplasmic form consisting of NFkappaB bound to IkappaB. NFkappaB was originally identified as a heterodimeric DNA binding protein complex consisting of p65 (RelA) and p50 (NFKB1) subunits. Other identified subunits include p52 (NFKB2), c-Rel, and RelB. The p65, cRel, and RelB subunits are responsible for transactivation. The p50 and p52 subunits possess DNA binding activity but limited ability to transactivate. p52 has been reported to form transcriptionally active heterodimers with the NFkappaB subunit p65, similar to p50/p65 heterodimers. The heterodimers of p52/p65 and p50/p65 are regulated by physical inactivation in the cytoplasm by IkappaBalpha. IkappaBalpha-??binds to the p65 subunit, preventing nuclear localization and DNA binding. Low levels of p52 and p50 homodimers can also exist in cells.
NFkB p65 (Rel A) peptide corresponding to a region near the C-terminus of the human protein conjugated to Keyhole Limpet Hemocyanin (KLH).
Immunogen Type
Storage Condition
Store vial at -20° C or below prior to opening. This vial contains a relatively low volume of reagent (25 µL). To minimize loss of volume dilute 1:10 by adding 225 µL of the buffer stated above directly to the vial. Recap, mix thoroughly and briefly centrifuge to collect the volume at the bottom of the vial. Use this intermediate dilution when calculating final dilutions as recommended below. Store the vial at -20°C or below after dilution. Avoid cycles of freezing and thawing.
Application Note
This protein A purified mouse monoclonal antibody is directed against NFkB p65 (Rel A) and a 1:1000 dilution recognizes a 65 kD band by Western blot against HeLa whole cell lysate. Control peptide (100-4165p) is sold separately.
This product was purified from concentrated tissue culture supernate by Protein A chromatography and showed a single band by IEP (immunoelectrophoresis) when tested with anti-mouse antibody. Reactivity was confirmed by ELISA against peptide conjugated carrier protein and by Western blot against HeLa whole cell lysate.
Disclaimer Note-General
This product is for research use only and is not intended for therapeutic or diagnostic applications. Please contact a technical service representative for more information. All products of animal origin manufactured by Rockland Immunochemicals are derived from starting materials of North American origin. Collection was performed in United States Department of Agriculture (USDA) inspected facilities and all materials have been inspected and certified to be free of disease and suitable for exportation. All properties listed are typical characteristics and are not specifications. All suggestions and data are offered in good faith but without guarantee as conditions and methods of use of our products are beyond our control. All claims must be made within 30 days following the date of delivery. The prospective user must determine the suitability of our materials before adopting them on a commercial scale. Suggested uses of our products are not recommendations to use our products in violation of any patent or as a license under any patent of Rockland Immunochemicals, Inc. If you require a commercial license to use this material and do not have one, then return this material, unopened to: Rockland Inc., P.O. BOX 5199, Limerick, Pennsylvania, USA.
General Reference
Unlap, M.T. and Jope, R.S. (1997). Dexamethasone attenuates NF-kappaB DNA binding activity without inducing IkappaB levels in rat brain in vivo. Molecular Brain Research 45: 83-89. Poser, I., Golob, M., Buettner, R. and Bosserhoff, A.K. (2003) Upregulation of HMG1 Leads to Melanoma Inhibitory Activity Expression in Malignant Melanoma Cells and Contributes to Their Malignancy Phenotype. Mol. Cell Biol. 23(8): 2991-2998. Jobin, C, et al. (1999) Curcumin blocks cytokine-mediated NF-kappa B activation and proinflammatory gene expression by inhibiting inhibitory factor I-kappa B kinase activity. J. Immunol. 163(6): 3474-3483.
Specific Reference
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Product Type
Primary Antibodies;
Reacts With
human, mouse, rat, chicken
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Product Type
Primary Antibodies;
Reacts With
human, mouse, rat
Catalog Number

Product Type
Primary Antibodies;
Reacts With
Catalog Number

Product Type
Proteins and Peptides;
Catalog Number

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Conjugation Reference
Molecular Weight
Excitation Wavelength
Conjugation Chemistry