PART I:EXPERIMENTS Experiment 1 Protein concentration determination According to the proteins'physical,chemical or biological properties,we have developed lots of methods to determining protein concentration.In this article we will study the principle of several protein determination methods to understand their protocols and application. The Lowry method for protein quantitation 1.Principle td is based on both the Biuret reaction.n which the peptide bonds of to prod which 11B0010-u10.J111p with coppe reacts but in essence phosphomolybdotungstate is reduced to heteropolymolybdenum blue by the copper-catalyzed oxidation of aromatic amino acids.The reactions result in a strong blue color,which depends partly on the tyrosine and tryptophan content. Sensitivity of the procedure of Lowry et al.is moderately constant from protein to protein,and it has been so widely used that Lowry protein estimations are a completely acceptable alternative to a rigorous absolute determination in almost all circumstances in which protein mixtures or crude extracts are involved.The method is sensitive down to about 0.01 mg of protein/mL,and is best used on solutions with concentrations in the range 0.01-1.0 mg/mL of protein 2.Materials 1.Standards:Use a stock solution of standard protein (e.g.bovine serum albumin fraction V)containing 250 ug/mL protein in distilled water,stored frozen at _20℃ 2.Complex-forming reagent:Prepare immediately before use by mixing the following stock solutions in the proportion 50:50:1:1 (by vol).(first mix A and B.C and D,respectively,and then mix the two solutions): Solution A:4%(w/v)NazCO;in distilled water Solution B:0.2 mol/L NaOH in distilled water olutio C. 1%(w/v)CuSO5H2O in distilled water Solution D:2%(w/v)sodium potassium tartrate in distilled water. 3.Folin reagent(commercially available):Use at 1 M concentration. 4.Sample:human serum diluted 100 folds with distilled water,store frozen at -20℃. 4