Review for Mouse Anti-Flag Antibody

Protein extract from mouse MEFcells was prepared in radioimmunoprecipitation assay lysis buffer (sc-24948; Santa Cruz Biotechnology) supplied with PMSF, protease inhibitor mixture, sodium orthovanadate, or RNase inhibitor. Lysates were quantified using a Bio-Rad protein assay kit. Forty micrograms of proteins were subjected to SDS-PAGE and transferred onto nitrocellulose membranes and blocked with 5% non-fat milk in TBS-T (20 mMTris, 500 mMNaCl, and 0.1% Tween 20) at room temperature for 1 h with rocking. The membranes were probed with primary Antibody (1/1000 dilution) overnight at 4°C. After washing with TBS-T, the membranes were incubated with HRP-conjugated secondary Antibodies (Thermo Scientific, Rockford, IL) in 5% non-fat milk/TBS-T at room temperature for 1 h. The protein–Antibody complex was detected by ECL substrate (cat # 32106; Thermo Scientific).

Research published: Yang S, Deng P, Zhu Z, Zhu J, Wang G, Zhang L, Chen AF, Wang T, Sarkar SN, Billiar TR, Wang Q. 2014. Adenosine deaminase acting on RNA 1 limits RIG-I RNA detection and suppresses IFN production responding to viral and endogenous RNAs. J Immunol,193: 3436-3445"