Immunofluorescence was performed on fixed and permeabilized Jurkat cells treated with Calyculin A (50 ng/ml/20min) for detection of NF-kappa-B p65 (pS 536) using Anti-NF-kappa-B p65 [pS536] Rabbit Polyclonal Antibody (Product# 720006, 2 ug/ml) and labeled with Goat anti-Rabbit IgG (H+L) Superclonal™ Secondary Antibody, Alexa Fluor® 488 conjugate (Product # A27034, 0.4 ug/ml, 1:2500). Panel a) shows representative cells that were stained for detection and localization of NF-kappa-B p65 [pS536] protein (green), Panel b) is stained for nuclei (blue) using SlowFade® Gold Antifade Mountant with DAPI (Product # S36938, 1:50). Panel c) represents cytoskeletal F-actin staining using Alexa Fluor® 594 Phalloidin (Product # A12381, 1:200). Panel d) is a composite image of Panels a, b and c clearly demonstrating nuclear localization of NF-kappa-B p65 (pS 536). Panel e) represents control cells with no primary antibody to assess background.
|Tested species reactivity||Human|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Phosphopeptide against Human NF-kappa-B p65 (aa 531-539)|
|Storage buffer||PBS, pH 7.2, with 30% glycerol, 0.1% BSA|
|Contains||0.09% sodium azide|
|Storage Conditions||Store at 4°C short term. For long term storage, store at -20°C, avoiding freeze/thaw cycles.|
|Tested Applications||Dilution *|
|Flow Cytometry (Flow)||2-4 µg/10^6 cells|
|Immunocytochemistry (ICC)||2 µg/ml|
|Immunofluorescence (IF)||2 µg/ml|
|Western Blot (WB)||2-3 µg/ml|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
This antibody is predicted to react with Monkey, Rat, Mouse, Pig, Bovine, Rabbit and Dog.
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.