Protocol for Anti-SNAP-tag® Antibody (Polyclonal) (P9310)

Protocol

1. Perform SDS-PAGE and protein transfer according to standard Western blotting methods.
2. After protein transfer, wash membrane with 25ml of 1X TBS for 5 minutes at room temperature.
3. Incubate membrane in 25ml of Blocking Buffer for 1 hour at room temperature.
4. Wash membrane 3 times for 5 minutes each with 15ml of TBST Wash Buffer.
5. Incubate membrane with Anti-SNAP-tag® Antibody (NEB #P9310S) at a 1:1000 dilution in 20ml of Blocking Buffer with gentle rocking overnight at 4ºC or for 1 hour at room temperature.
6. Wash membrane 3 times for 5 minutes each with 15ml of TBST Wash Buffer.
7. Incubate membrane with a commercially available secondary antibody (e.g. Anti-Rabbit IgG HRP-linked antibody) in 20ml of Blocking Buffer with gentle rocking for 1 hour at room temperature. Use the working dilution suggested by the manufacturer of the secondary antibody.
8. Wash membrane 3 times for 5 minutes each with 15ml of TBST Wash Buffer.
9. Proceed to chemiluminsecent detection of protein bands with a commercially available detection reagent.