Fast Activated Cell-based ELISA (FACE™) Kits provide a simple, sensitive method for detecting protein phosphorylation directly in the cell, without making extracts or performing electrophoresis and membrane blotting. These 96-well, high-throughput assays are available in both colorimetric and chemiluminescent formats for over 20 different targets (see list at right). For complete details, click the FACE™ Method tab below.
FACE NF?B p65 Profiler Kits provide 96 rxns each of 3 antibodies that enable you to monitor and compare the levels of NF?B p65 phosphorylated at two different sites, as well as total NF?B p65. The phospho-NF?B p65 (S468) antibody specifically recognizes NF?B p65 when phosphorylated at Ser468, while phospho-NF?B p65 (S536) antibody specifically recognizes NF?B p65 that is phosphorylated at Ser536. The total-NF?B p65 antibody recognizes NF?B p65 regardless of its phosphorylation state. Click the NF?B p65 Info tab below for data and more information.
Figure 1: Measurement of phosphorylated and total NF?B p65.
HeLa cells were cultured in 96-well plates and serum-starved for 16 hours. Cells were then treated with 20 ng/ml TNF-? and 50 nM Calyculin A for 15 minutes and fixed. Total and phospho NF?B p65 were each assayed in triplicate using the phospho and total NF?B p65 antibodies included in the FACE NF?B p65 Profiler Kit. Data was plotted after correction for cell number (performed through use of Crystal Violet).
The phospho-NF?B p65 (S468) was raised against a synthetic phospho-peptide corresponding to residues surrounding Serine 468 of human NF?B p65 and does not cross-react with other sites. The phospho-NF?B p65 (S536) was raised against a synthetic phospho-peptide corresponding to residues surrounding Serine 536 of human NF?B p65 and does not cross-react with other sites. The total-NF?B p65 antibody recognizes NF?B p65 proteins regardless of the phosphorylation state.
NF?B p65 Overview
The transcription factor NF?B (NF-kappa-B p65, RelA, nuclear factor ?B) is a key component for the inducible expression of a wide va