Antibody data
- Antibody Data
- Antigen structure
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- Validations
- Western blot [2]
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Validation data
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- Product number
- 44-963G - Provider product page
- Provider
- Invitrogen Antibodies
- Product name
- Phospho-PKC beta-1 (Thr642) Polyclonal Antibody
- Antibody type
- Polyclonal
- Antigen
- Synthetic peptide
- Reactivity
- Human
- Host
- Rabbit
- Isotype
- IgG
- Vial size
- 100 µL
- Storage
- -20°C
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Supportive validation
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Peptide Competition and Phosphatase Treatment. Lysates prepared from K562 cells stimulated with PMA were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were either left untreated (1-4) or treated with lambda phosphatase (5), blocked with a 5% BSA-TBST buffer for one hour at room temperature, and incubated with PKCbetaI (pT642) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 5), the non-phosphopeptide corresponding to the immunogen (2), a generic phosphothreonine-containing peptide (3), or, the phosphopeptide immunogen (4). 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 PKCbetaI (pT642) blocks the antibody signal. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific.
- Submitted by
- Invitrogen Antibodies (provider)
- Main image
- Experimental details
- Peptide Competition and Phosphatase Treatment. Lysates prepared from K562 cells stimulated with PMA were resolved by SDS-PAGE on a 10% polyacrylamide gel and transferred to PVDF. Membranes were either left untreated (1-4) or treated with lambda phosphatase (5), blocked with a 5% BSA-TBST buffer for one hour at room temperature, and incubated with PKCbetaI (pT642) antibody for two hours at room temperature in a 3% BSA-TBST buffer, following prior incubation with: no peptide (1, 5), the non-phosphopeptide corresponding to the immunogen (2), a generic phosphothreonine-containing peptide (3), or, the phosphopeptide immunogen (4). 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 PKCbetaI (pT642) blocks the antibody signal. The data also show that phosphatase stripping eliminates the signal, verifying that the antibody is phospho-specific.