双向电泳试剂配制以及protocol.doc

水化液 8mM Urea12g4.82%CHAPS0.5g0.20.5%IPG Buffer125ul500.002%Bromophend Blue50ul (1%BB)2.0DD H2OTo25mlTo 10ml 分装成500ul每管，冻于20，用前融化。SDS平衡液123Tris-HCL PH8.8(F液)0.54ml1.081.8Urea（Fw60.06）5.41g10.8218.2Glycerol (87%V/V)4.53g9.0615.1SDS0.3g0.61.0Bromophenol Blue30ul60100DDH2OTo15mlTo30To50第一天配制2管，摇晃过夜第二天用前，一管加入DTT（100mg/10ml） 另一管加入碘乙酰胺（250mg/10ml）碘乙酰胺较难溶，多摇晃几下II向SDS胶123Monomer Solution(E液)50.04ml83.4125.14Vesoling gel buffer(F液)30ml507510%SDS(G液)1.2ml23DDH2O38.16ml63.695.410%Ammonium Peosulfate (0.486-25G)600ul1ml1.5TEMED (10%AP:0.151.5ml)39.6ul6699E液Acrylumicle 30%150gBis- Acylumicle 0.8%4gDDH2OTo 500ml4贮藏F液Tris-base 1.5m181.7gDDH2OTo 750mlHCLTo PH8.8DDH2OTo 1L4贮藏G液SDS 10% (W/V)5gDDH2OTo 50ml室温二向下槽液25mM TrisHCL PH 8.312.12g192mM gly57.6g0.1% SDS4.5g定容至500ml（DDH2O），搅拌，移至4L试剂瓶，再加入3.5L DDH2O，需前一天加入电泳槽，以消泡。琼脂葡糖液二向下槽液100mlAgarose(NA or M) 0.5%0.5gBromophenol 0.002%200ul87溶解，室温保存1%溴酚蓝液Bromophend Blue 1%100mgTris-base60mgDDH2OTo 10ml室温保存固定液（无水）乙醇150ml乙酸37.5mlDDH2O188ml敏化液123(无水)乙醇112.5ml225337.5Sodium thiosulfate0.75g1.52.25Sodium acetate (17%)25.5g5176.5DDH2OTo 350mlTo700To1050银染液123AgNO3 (2.5%)0.9375g1.8752.8125DDH2OTo 375 mlTo750To1125显色液123Sodiun carbonate(6.26g)9.375g18.7528.125甲醛（37%）用前加150ul300450DDH2OTo 375 mlTo750To1125终止液123EDTA2Na（365g）5.475g10.9516.425DDH2OTo 375mlTo750To1125