Wash and dry the chamber. Assemble the chamber. Don't forget to place a piece of membrane between the two halves of the chamber. Inject 1 ml of sample and control in each side of the chamber. Stir the chamber for 8 hours (or O/N) in the cold room. Take the sample out of the chamber and count the cpm. Sample preparation: cAR1; 1 mg/mL BSA; 10% sucrose; 100 mM NaCl; 0.1% DM; 40 mM Tris; pH 7.9 Hot cAMP; Add 10 mM DTT for CHIFF binding. Don’t add DTT for pure cAR1 binding. Control: No cAR1 added; the rest similar as described above.
This page was last edited 06/26/2003 |