Anti-Btk (phospho Y223) Antibody

The reagent is a rabbit polyclonal IgG raised against Bruton's tyrosine kinase (Btk) phosphorylated at tyrosine 223, a Tec family kinase modification that marks the catalytically active state following B-cell receptor (BCR) engagement. Supplied by Antibodies.com as 200 µl of unconjugated liquid, it is validated for Western blot and immunofluorescence across human, mouse, and rat samples.

Y223 sits within the Btk SH3 domain and is generated through trans-autophosphorylation after Src-family kinases (Lyn, Syk) phosphorylate the activation-loop residue Y551. Phospho-Y223 reaches maximal signal within roughly five minutes of anti-IgM stimulation and decays toward baseline by thirty minutes, making the marker a precise temporal readout of receptor-proximal kinase activity. This brief kinetic window is why the antibody is widely used in B-lymphoma lines (Raji, Ramos, TMD8) and in primary B cells to confirm target engagement of covalent BTK inhibitors (ibrutinib, acalabrutinib, zanubrutinib) in dose-response and washout studies.

For Western blot, the antibody is best resolved on 8% polyacrylamide gels and paired with a total Btk antibody for normalization, while the unconjugated format supports custom HRP, biotin, or fluorophore conjugation at the bench. Immunofluorescence on suspension B cells calls for gentle paraformaldehyde fixation to preserve the phospho-epitope, since over-fixation erodes signal. As a research use only reagent, store refrigerated at 4 °C, protected from light, with aliquots prepared to limit freeze-thaw cycles across multiple uses; sodium azide is typically added as a preservative.