- Proteins and Peptides
- Lysates and Cell Lines
Western Blot Procedure
1) Wet Nitrocellulose membrane with PBS + NP40 [PN].
2) Pour off PN.
3) Dilute anti-RNP M3/M4 (catalog# NB 200-315) in 5%NFDM + NP40.
4) Incubate the primary antibody with the membrane for 1 hour, at room temperature (RT), gently rocking.
5) Wash 1x with PN for 10 minutes. Wash 2x with PN for 5 minutes, each.
6) Dilute anti-mouse-HRP (Amersham) in 5%NFDM + NP40 at 1:5,000.
7) Incubate the secondary antibody with the membrane for 45 minutes, at RT, gently rocking.
8) Wash 1x with PN for 10 minutes. Wash 2x with PN for 5 minutes, each.
9) Wash 1x with PBS for 5 minutes.
10) Develop using Amersham ECL components.
**NOTE: HeLa nuclear cell extracts can be used as a positive control for this antibody.