Western blot analysis was performed on whole cell extracts (30 ug lysate) of HeLa (Lane 1), NIH/3T3 (Lane 2), A-431 (Lane 3), K-562 (Lane 4), MCF7 (Lane 5), Jurkat (Lane 6), THP-1 (Lane 7) and HCT 116 (Lane 8). The blot was probed with Anti-NFkB p52 Mouse Monoclonal Antibody (Product # MA5-17217, 1 ug/ml) and detected by chemiluminescence using Goat anti-Mouse IgG (H+L) Superclonal™ Secondary Antibody, HRP conjugate (Product # A28177, 0.4 ug/ml, 1:2500 dilution). A 52 kDa band corresponding to NFkB p52 was observed across the cell lines tested. Known quantity of protein samples were electrophoresed using Novex® NuPAGE® 4-12 % Bis-Tris gel (Product # NP0321BOX), XCell SureLock™ Electrophoresis System (Product # EI0002) and Novex® Sharp Pre-Stained Protein Standard (Product # LC5800). Resolved proteins were then transferred onto a nitrocellulose membrane with iBlot® 2 Dry Blotting System (Product # IB21001). The membrane was probed with the relevant primary and secondary Antibody following blocking with 5 % skimmed milk. Chemiluminescent detection was performed using Novex® ECL Chemiluminescent Substrate Reagent Kit (Product # WP20005).
|Tested species reactivity||Human, Mouse, Rat|
|Host / Isotype||Mouse / IgG2b|
|Immunogen||Recombinant human protein purified from E.coli (His-NFkB 1-454 a.a.)|
|Storage buffer||HEPES with 0.15M NaCl, 0.01% BSA, 50% glycerol|
|Contains||0.03% sodium azide|
|Tested Applications||Dilution *|
|ELISA (ELISA)||Assay Dependent|
|Flow Cytometry (Flow)||3-5 µg/10^6 cells|
|Western Blot (WB)||0.5-2 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.
A suggested positive control for this product is K562 cells.
NF-kappaB (Nuclear Factor kappa B) is a nuclear transcription factor found in all cell types and is involved in cellular responses to stimuli such as stress, cytokines, free radicals, ultraviolet irradiation, and bacterial or viral antigens. NF-kappaB plays a key role in regulating the immune response to infection. Consistent with this role, incorrect regulation of NF-kappaB has been linked to cancer, inflammatory and autoimmune diseases, septic shock, viral infection and improper immune development. There are five members in the NF-kappaB family: NF-kappaB1, NF-kappaB2, RelA (also named p65), RelB, and c-Rel. The most common form of NF-kappaB is the p50/RelA heterodimer, although other forms of NF-kappaB dimers, such as p50/p50, p52/p52, p52/RelA, p50/c-Rel, c-Rel/c-Rel, p52/RelB, and p50/RelB, have also been identified in some types of cells. The primary role of NF-kappaB is to maintain normal cellular functions that range from cell-to-cell communication to cell motility, cell cycle progression, and cell lineage development. The activity of NF-kappaB is tightly regulated by interaction with inhibitory IkappaB proteins.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.