IP-蛋白质印迹:

IP/WB analysis of active Cdc42 protein. Purified recombinant Cdc42 proteins were loaded with GDP (lane 1) or GTPγS (lane 2). These proteins were immunoprecipitated with anti-Cdc42-GTP mouse monoclonal antibody (Cat. # 26905). After SDS/PAGE, the membrane filter was probed with anti-Cdc42 mouse monoclonal antibody (Cat. # 26008).

免疫荧光:

Immunofluorescent labeling of Cdc42-GTP in MS1 mouse endothelial cells. The MS1 cells cultured on glass coverslips (Fisher) in DMEM containing 5% FBS were fixed with 4% paraformaldehyde for 10 minutes at RT (room temperature) and permeabilized in PBSN (PBS+0.1%NP40) for 15 minutes at RT. The cells were stained with primary antibody (Cdc42-GTP) at 1: 25 dilution in Invitrogen’s CAS-BLOCK (00-8120) reagent at RT for 1 hour. After brief washing in PBSN (3x5min), a secondary Alexa555 conjugated goat anti mouse antibody (Invitrogen A21424) was applied in 1: 200 in CAS-BLOCK at RT for 1 hour. The coverslips were then washed in PBSN (3x5min) and mounted using vectorsheld’s hard set mounting medium (H-1500), and examined using a Zeiss inverted fluorescence microscope.