Comparison of the Lowry and Coomassie Blue methods for the determination of protein concentration.
1. The Lowry and Coomassie Blue methods were compared for precision when applied to Schistosoma mansoni proteins, using bovine serum albumin (BSA) as standard. 2. The absorbance of five different concentrations was measured and the experiments were run in parallel to reduce bias in the comparison. Thus, for a given technique, all photometric data were obtained from reaction mixtures (BSA and test solutions) prepared and analyzed as simultaneously as possible. 3. To interpret the results, polynomial functions of different degrees--up to the third--were calculated to fit the absorbance values to the respective protein concentrations of BSA or worm protein present in the reaction mixtures, and the standard errors of all slopes were also calculated. 4. The protein concentration of worm extracts was calculated by two methods: 1) the ratio of the first-degree slopes of the polynomials applied to absorbance = f(x), with x being either mg BSA or ml worm extract, and 2) the utilization of the BSA function parameters to convert any worm absorbance value to protein concentration. 5. The results obtained with the slope ratio method were variable. However, the function method seemed to be reliable, especially when applied to Coomassie Blue data. When the results were derived from the BSA cubic function the concentration of soluble worm protein was calculated with a coefficient of variation of 1.20%.
Vieira
E G
EG
Laboratório Especial de Biologia Molecular, Instituto Butantan, São Paulo, Brasil.
Henriques
S B
SB
D000818:Animals; D005069:Evaluation Studies as Topic; D015801:Helminth Proteins / Q000032:analysis*; D007202:Indicators and Reagents; D008433:Mathematics; D008982:Molybdenum; D012394:Rosaniline Dyes; D012550:Schistosoma mansoni / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000032:analysis; D012996:Solutions; D013053:Spectrophotometry / Q000379:methods*; D017973:Tungsten Compounds
C029556:Folin's phenol reagent; D015801:Helminth Proteins; D007202:Indicators and Reagents; D012394:Rosaniline Dyes; D012996:Solutions; D017973:Tungsten Compounds; D012710:Serum Albumin, Bovine; C048139:Coomassie blue; D008982:Molybdenum