Jurkat cells were treated for 30 min with 100 ug/mL ALLN (N-Acetyl-Leu-Leu-Norleucinal; a Calpain inhibitor and also proteasome inhibitor that prevents IkBa dephosphorylation) followed by incubation with (lanes 2 & 4) or without 1 nM TNF-a (1 & 3). The membranes were blotted with NFKBIA monoclonal antibody, clone 39A1413 (Cat # MAB0056) (lanes 1 & 2) or NFKBIA monoclonal antibody, clone 6A920 (Cat # MAB0057) (that recognizes both non-phospho and phosphorylated forms of IkBa) (lanes 3 & 4) and immunoreactivity was detected by ECL. The data shows that Cat # MAB0056 detects specifically the phosphorylated form of NFKBIA.
Western Blot (Cell lysate)
Western blot analysis of NFKBIA. Using NFKBIA monoclonal antibody, clone 6A920 (Cat # MAB0057) at 2 ug/mL in (A) Daudi and (B) NIH/3T3 whole cell lysate.
NFKB1 (MIM 164011) or NFKB2 (MIM 164012) is bound to REL (MIM 164910), RELA (MIM 164014), or RELB (MIM 604758) to form the NFKB complex. The NFKB complex is inhibited by I-kappa-B proteins (NFKBIA or NFKBIB, MIM 604495), which inactivate NF-kappa-B by trapping it in the cytoplasm. Phosphorylation of serine residues on the I-kappa-B proteins by kinases (IKBKA, MIM 600664, or IKBKB, MIM 603258) marks them for destruction via the ubiquitination pathway, thereby allowing activation of the NF-kappa-B complex. Activated NFKB complex translocates into the nucleus and binds DNA at kappa-B-binding motifs such as 5-prime GGGRNNYYCC 3-prime or 5-prime HGGARNYYCC 3-prime (where H is A, C, or T; R is an A or G purine; and Y is a C or T pyrimidine).[supplied by OMIM
IkappaBalpha,nuclear factor of kappa light chain gene enhancer in B-cells