Detection of total NF-kappaB p65, using ab119637 on HeLa cells stimulated with various concentrations of TNF alpha.
Using the PhosphoTracer Total NF-kappaB p65 assay, the amount of lysate required to detect NF-kappaB p65 in TNFalpha-treated HeLa cell lysates was examined. Up to 50 μg of lysate/well was loaded into replicate wells of a PhosphoTracer microplate, and analysed for total NF-kappaB p65. Total NF-kappaB p65 was readily detected in less than 10 μg of lysate/well.
Using the NF-kappaB p65 assay kits, a significant stimulation of NF-kappaB p65 phosphorylation at Ser536 is detected in HeLa cells treated with TNFalpha for 10 minutes (+) compared with untreated cells (-), while no change in total NF-kappaB p65 levels is observed.
Using Western blot, a significant stimulation of NF-kappaB p65 phosphorylation at Ser536 is detected in HeLa cells treated with TNFalpha for 10 minutes (+) compared with untreated cells (-), while no change in total NF-kappaB p65 levels is observed.