- Proteins and Peptides
- Lysates and Cell Lines
1. Proteins are separated on a 10% SDS-PAGE gel.
2. Proteins are transferred to Immobilon-P membranes (Millipore Co.).
3. Following the protein transfer, the membrane is blocked with PBS-T [PBS + 0.07% Tween-20]+ 5% NFDM.
4. Anti-RNase L [cat# NB 100-351] is diluted 1:1,000, in blocking buffer and incubated for 2 hour at room temperature, gently shaking.
5. The membrane is then washed, 3 times with PBS-T, 5 minutes each.
6. Secondary antibody is incubated for 1 hour at room temperature, gently shaking.
7. The membrane is then washed, 4 times with PBS-T, 5 minutes each.
8. Membrane is developed using ECL reagents.
NOTE: Cell extracts of insect cells expressing human RNase L (0.1 ug) and Hey 1B (100 ug) were used as positive controls for this antibody.