This BIK (phospho Thr33) Cell Based ELISA Kit allows for the detection of BIK (phospho Thr33) and the effects that certain stimulation conditions have on BIK (phospho Thr33) expression in different cell lines. Qualitative determination of BIK (phospho Thr33) concentration is achieved by an indirect ELISA format. In essence, the BIK (phospho Thr33) is captured by Anti-BIK (phospho Thr33) Antibodies which in turn are detected by HRP-conjugated secondary antibodies. Through this binding, the HRP enzyme (conjugated to the secondary antibody) can catalyze a colorimetric reaction upon substrate addition. Due to the qualitative nature of this BIK (phospho Thr33) Cell Based ELISA Kit, multiple normalization methods are described: 1) Anti-GAPDH Antibody is included to serve as an internal positive control in normalizing the target absorbance values. 2) Following the colorimetric measurement of HRP activity via substrate addition, the Crystal Violet whole-cell staining method is used to determine cell density. After staining, the results can be analyzed by normalizing the absorbance values to cell amounts, by which the plating difference can be adjusted. 3) Anti-BIK Antibody is provided for normalization purposes. The absorbance values obtained for the non-phosphorylated target can be used to normalize the absorbance values for the phosphorylated target.
Figure 4: Western Blot - BIK (phospho Thr33) Cell Based ELISA Kit (A102761)
The mouse monoclonal antibody to GAPDH is used as a positive control inBIK (phospho Thr33) Cell Based ELISA Kit. Anti-GAPDH Antibody was tested for specificity by western blot with tissue lysates from human, mouse, and rat.
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