44-232G

antibody from Invitrogen Antibodies

Targeting: ZAP70 SRK, STD, ZAP-70

Western blot (Supportive data in Antibodypedia)

Antibody Data

Product number

44-232G

Provider

Invitrogen Antibodies

Product name

Phospho-Zap-70 (Tyr315, Tyr319) Polyclonal Antibody

Antibody type

Polyclonal

Antigen

Synthetic peptide

Reactivity

Human

Host

Rabbit

Isotype

IgG

Vial size

100 µL

Storage

-20°C

Western blot

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Peptide Competition and Phosphatase Treatment. Lysates prepared from Jurkat cells left untreated (1) or treated with H2O2 (2-6) were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were either left untreated (1-6) or treated with YOP phosphatase (7), blocked with a 5% BSA-TBST buffer for one hour at room temperature, and incubated with ZAP-70 (pYpY315/319) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 2, 6), the non-phosphopeptide corresponding to the immunogen (3), a generic phosphotyrosine-containing peptide (4) the phosphopeptide immunogen (5) or, the phosphopeptide corresponding to ZAP-70 (pY315) (6). After washing, membranes were incubated with goat F (ab’)2 anti-rabbit IgG HRP conjugate (Product # ALI4404) and bands were detected using the Pierce SuperSignal™ method. The data show that only the peptide corresponding to ZAP-70 (pYpY315/319) completely blocks the antibody signal. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific.