[Detection of nuclear factor kappa B by immunochemical staining and image analysis].

Objective: To find a relatively simple and safe method for quantitative assay of nuclear factor (NF)-kappa B activity.

Methods: Immunochemical staining of NF-kappa B/p65 subunit with its antibody was performed in human umbilical vein endothelial cells stimulated with advanced glycation end products-modified human serum albumin (AGE-HSA). The ratio of p65 subunit staining in the nuclei and cytoplasm was determined by means of imaging analysis, and the results were compared with those of electrophoretic mobility shift assay (EMSA).

Results: NF-kappa B could be activated by AGE-HSA in a time- and dose-dependent manner, and the results obtained from immunochemical staining were consistent with those from EMSA.

Conclusion: p65 immunochemical staining and subsequent image analysis is feasible in the quantitative detection of NF-kappa B activation.