Antibody From Cell Signaling Technology for Decent Mouse Cyclin E1 detection.

Results Summary

Cell lines from syngeneic mouse models were over-expressed Cyclin E1. The immunoblotting procedure is described below. Lyse cells by adding RIPA (with Proteinase and Phosphatase inhibitors) buffer. Heat 20µg of protein to 95–100°C for 5 min. Load 20µg of protein onto SDS-PAGE gel. Electrotransfer to PVDF membrane (Semi-dry, 20V, 1 and half hours). Incubate membrane in blocking buffer (1X TBST with 5% nonfat dry milk) for 1 hour at room temperature. Incubate membrane with diluted (1:1,000) primary antibody in 5% BSA, 1X TBS, 0.1% Tween20 at 4°C with gentle shaking, overnight. Incubate membrane with the Rabbit-specific HRP-conjugated secondary antibody in blocking buffer with gentle agitation for 1 hour at room temperature.