蛋白质测定.doc

Cu2+540nm0.5g/L10g/LTrisGood10%1m?NaOHLowryCu+-20mg/L400mg/LpHpH10Cu2+-Cu2+-280nm280nm0.10.5mg/ml260nm280nm1260nm10.1%280nm0.52.5230nm0.1%225nm215nm5.011.7280nm0.58230nm215nm225nm10l/ml100l/ml pHABradford()(Biuret)Folin?(Lowry)1976Bradford(Bradford)G-250(lmax)465nm595nm()595nmA595Bradford:(1)Lowry1mg-Lowry(2)521520Lowry(3)LowryK+Na+Mg2+TrisEDTA:(1)Bradford g?(2):Triton X-100(SDS)0.1NNaOH(0.1NLowary)(3)Beer()1. :(1) g?(BSA)1.0mg/ml0.1mg/ml(2)G-250:100mgG-25050ml 95%120ml 85%12. :(1) (2) (3)16()1. (1)16131.0mg/ml:00.010.020.040.060.080.1ml0.1ml5.0mlG-250()8910(2)25595nmA59510.1mlH2O5.0mlG-250:()95%(3)(mg)A595A5950.5mg/mlA5950.502. (10-100mg/ml)()0.5ml1.0ml0.5ml1.0ml H2OG-2505.0ml595nm0.05mg/mlA595.Bouguer1729Lambert1760:T=I/I0=e-kbI0Iekb(cm)BouguerBeer-Bouguer:T=I/I0=e-kbcc(g/Lmg/L)10:A=-logT=-log(I/I0)=log(I0/I)=bcA=BeerA=kbccg/LbcmKa/L/(gcm,A=kbcA=abcA=kbc1mol/L,b1cmk/L/(molcmA=kbcA=c1mol/LNADH260nm15000260NADH=15103340nm6220340NADH=6.22103A=kbccw/vbcmkE%A=kbcA=E%bcaEB . 280nm238nmNH42SO4pHpHpHpH1. 280nm280nm280nm280nmA2800.11.0mg/ml1cm1mg/mlA2801.0A11cmA11cm1%1cmA11cm, = (A28010 )/ A11cm,280nm (mg/ml)(Q 1%?10mg/ml)2. 280nm260nm280nm10260nm260nm260nm280nm2280nm260nmA280/A260 ? 1.8 A280/A260 ? 0.5A280A260 =1.45A2800.74A260 (mg/ml)3. 215nm225nm280nm215nm225nm20100 mg/ml5.0ml215nm225nm D= A215 A225D20100mg/mlNaClNH42SO40.1MTris0.1