Rockland

NFkB p65 Monoclonal Antibody (27F9.G4)

Product Image Gallery

NFkB p65 Antibody in Western Blot (WB) — Rockland anti NFKB p65 (Rel A) monoclonal antibody 200-301-065 was used to detect ~65 kDa 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.

NFkB p65 Antibody in Immunohistochemistry (IHC) — Rockland Antibody 200-301-065 has been tested in immunohistochemistry, analyzed by an anatomic pathologist and validated for use in IHC applications against formalin-fixed, paraffin-embedded human tissues. showed moderate to strong staining within many tissues, including epithelium of the breast, small intestine, kidney, pancreas, prostate, skin, placenta, and uterus, as well as within neurons and lymphoid tissues such as spleen, thymus, and tonsil. The antibody produced an excellent signal with almost no background staining at a concentration of 2.5 µg/ml. The image displayed shows specific staining in colon carcinoma as the precipitated red signal, with a hematoxylin purple nuclear counterstain. Image provided courtesy of LifeSpan Biosciences, Seattle, WA

NFkB p65 Antibody in Immunocytochemistry (ICC) — Anti NFkB monoclonal antibody - Immunocytochemistry. Tissue: Human Fibroblasts. Top: Before activation. Bottom: After activation with poly IC. Left: 600-401-271 Anti-p65 NLS specific - lot 18372. Right: 200-301-065 Anti-p65 C-Term monoclonal antibody - lot 26076. The two antibodies that are shown target different regions of the p65 protein. The different staining patterns are thought to correspond with different functional regions of the protein.

NFkB p65 Antibody in Immunofluorescence (IF) — 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 widefield, confocal and STED CW imaging on the same system.

NFkB p65 Antibody — PAGE-MAP (microsphere affinity proteomics) of Mouse Anti-NFKB p65 (Rel A) Antibody. (Catalog Number: 200-301-065, Lot Number: 26076). Antibody array western blot binding of gel free size separated fractions of multiple lysates (solid lines) and shotgun mass spectroscopy identification (dashed lines) of the target band run in parallel correlate confirming the specificity of this antibody against NFKB p65. Data was provided by the Lund-Johansen lab of Oslo University Hospital. For more information on PAGE-MAP/IP-MS identification of antibody specificity and its large-scale implementation for antibody validation see Sikorski et. al., (2018) Nature Methods 15, 909-912.

Product Details

Applications

Tested Dilution

Publications

Western Blot (WB)

1:1,000-1:5,000

Immunohistochemistry (IHC)

1:200-1:600

Immunohistochemistry (Paraffin) (IHC (P))

1:200-1:600

Immunocytochemistry (ICC)

Assay-dependent

Immunofluorescence (IF)

1:5,000

Immunoprecipitation (IP)

1:1,000-1:5,000

Product Specifications

Species Reactivity

Human

Host/Isotype

Mouse / IgG2a, kappa

Class

Monoclonal

Type

Antibody

Clone

27F9.G4

Immunogen

NFkB p65 (Rel A) peptide corresponding to a region near the C-terminus of the human protein conjugated to Keyhole Limpet Hemocyanin (KLH).

Conjugate

Unconjugated

Form

Liquid

Concentration

1 mg/mL

Purification

Protein A

Storage buffer

0.02M potassium phosphate, pH 7.2, with 0.15M NaCl

Contains

0.01% sodium azide

Storage conditions

-20° C, Avoid Freeze/Thaw Cycles

Product Specific Information

Store vial at -20° C prior to opening. Aliquot contents and freeze at -20° C or below for extended storage. Avoid cycles of freezing and thawing. Centrifuge product if not completely clear after standing at room temperature. This product is stable for several weeks at 4° C as an undiluted liquid. Dilute only prior to immediate use.

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.

Target Information

Nuclear factor kB p65 (NF-kB p65) is encoded by the RELA gene and is present on chromosome 11 in humans. NF-kB P65 is also known as RelA (v-rel avian reticuloendotheliosis viral oncogene homolog A) and belongs to the Rel family of proteins. It is one of the two subunits of NF-kB (Nuclear factor kappa-light-chain-enhancer of activated B cells) that heterodimerizes with the other subunits p50 or p52. However, binding of TNF alpha to its cognate receptor phosphorylates IKK which in turn phosphorylates I kB allowing proteasomal degradation of I kB. Phosphorylation of p65 at Ser 536 located in the carboxy-terminal transactivation domains (TAD) leads to nuclear localization of the transcriptionally active complex and NF-kB mediated transactivation of several downstream genes. It specifically plays a key role in transcription of immunoglobulin k (kappa) gene in mature B-lymphoid cells.

For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.

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Bioinformatics

Protein Aliases: MGC131774; NF KB; nf-kappa-b p65; NF-kappa-B p65delta3; NF-kappa-B transcription factor p65; NFkappaB p65; nfkb rela; NFkB-p50; NFKB1; nuclear factor; nuclear factor kappa b; Nuclear factor NF-kappa-B p65 subunit; Nuclear factor of kappa light polypeptide gene enhancer in B-cells 3; REL A; Transcription factor p65; v-rel avian reticuloendotheliosis viral oncogene homolog A; v-rel reticuloendotheliosis viral oncogene homolog A

Gene Aliases: NFKB3; p65; RELA

UniProt ID: (Human) Q04206

Entrez Gene ID: (Human) 5970

Molecular Function: nucleic acid binding transcription factor