Thiourea/urea lysis buffer
4% (w/v) CHAPS
2% (v/v) carrier ampholytes (pH 3-10)
10 mM Pefabloc proteinase inhibitor
To prepare thiourea/urea lysis buffer, dissolve 22.0 g of urea in deionized H2O, add 8.0 g of thiourea, and adjust the volume to 50 ml with deionized H2O. Add 0.5 g of Serdolit MB-1 mixed-bed ion-exchange resin (Serva), stir for 10 minutes, and filter. To 48 ml of the urea solution add 2.0 g of CHAPS, 1.0 ml of Pharmalyte (pH 3-10; Amersham Biosciences), 0.5 g of DTT, and, immediately before use, 50 mg of Pefabloc proteinase inhibitor (VWR).
