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Bradford protein assay
About: Bradford protein assay is a research topic. Over the lifetime, 635 publications have been published within this topic receiving 239107 citations.
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Patent•
27 Apr 2001
TL;DR: In this article, a method of protein precipitation, concentration and removal of non-protein agents from the protein solution was proposed, where protein solution is treated with a protein-precipitation agent containing an acidic agent, a salt and a precipitate forming agent.
Abstract: A method of protein precipitation, concentration and removal of non-protein agents from the protein solution wherein the protein solution is treated with a protein-precipitation agent containing an acidic agent, a salt and a precipitate forming agent. After precipitation, the protein precipitate is washed with a water miscible organic solvent agent to remove non-protein agents present in the protein precipitate.
29 citations
TL;DR: PPO in O. basilicum L. was extracted and purified through (NH4)2SO4 precipitation, dialysis, and a Sepharose 4B-l-tyrosine-p-aminobenzoic acid affinity column, and found that optimum temperature and pH were dependent on the substrates studied.
Abstract: A partial characterization of polyphenol oxidase (PPO) activity in Ocimum basilicum L. is described. PPO in O. basilicum L. was extracted and purified through (NH4)2SO4 precipitation, dialysis, and a Sepharose 4B-l-tyrosine-p-aminobenzoic acid affinity column. The samples obtained from (NH4)2SO4 precipitation and dialysis were used for the characterization of PPO. At the end of purification by affinity chromatography, 11.5-fold purification was achived. The purified enzyme exhibited a clear single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The molecular mass of the enzyme was estimated to be approximately 54 kDa. The contents of total phenolic and protein of O. basilicum L. extracts were determined. The total phenolic content of O. basilicum L. was determined spectrophotometrically according to the Folin-Ciocalteu procedure and was found to be 280 mg 100 g(-1) on a fresh weight basis. The protein content was determined according to the Bradford method. The enzyme showed activity to 4-methylcatechol, catechol, and pyrogallol substrates, but not to tyrosine. Therefore, of these three substrates, 4-methylcatecol was the best substrate due to the highest V(max)/K(m) value, followed by pyrogallol and catechol. The optimum pH was at 6, 8, and 9 for 4-methylcatechol, catechol, and pyrogallol, respectively. The enzyme had an optimum temperature of 20, 40, and 50 degrees C for 4-methylcatechol, catechol, and pyrogallol, respectively. It was found that optimum temperature and pH were dependent on the substrates studied. The enzyme activity with increasing temperature and inactivation time for 4-methylcatechol, catechol, and pyrogallol substrates decreased due to heat denaturation of the enzyme.
29 citations
TL;DR: In carbohydrate-rich samples, protein concentrations should be ascertained by precipitation from crude extracts and resolubilization in a noninterfering buffer, and sugar interference was much higher than expected from sugar molecules' individual contribution.
29 citations
TL;DR: A sensitive protein assay method which involves the reaction of TPPS4 with protein is described, and an absorption band with the maximum at 488 nm appears and the absorbance is proportional to the concentration of protein.
Abstract: A sensitive protein assay method which involves the reaction of TPPS4 with protein is described. When protein is added to TPPS4 solution, an absorption band with the maximum at 488 nm appears and the absorbance is proportional to the concentration of protein. Just Like the Soret absorption of the porphyrin, the new band is very narrow and there is no overlap at all between them, which means the free dyes would not give any background for the detection of the protein-TPPS4 complexes. A new spectrophotometric method for determination of protein has been constructed and applied to the determination of human plasma protein and urinary protein; The assay using microtiter plates has also been studied.
29 citations
TL;DR: Many proteins derived from the latex of Hevea brasiliensis that remain soluble in trichloroacetic acid (TCA) can be precipitated by phosphotungstic acid (PTA), but the use of PTA may not be fully compatible with the Bradford protein assay.
28 citations