protein estimation
TRANSCRIPT
• Determination of quantity of protein in given solution
• Colorimetric protein assay• Absorbance path length * Concentration• Absorbance Concentration (Path length:
Constant)
Biuret reaction + Reduction of Phosphomolybdate and Phosphotungstate from Folin Ciocateu’s (Phenol) reagent
• Reagent B : 1 : 5 Phenol reagent (phosphomolybdic-phosphotungstic acid)
10 ml of Folin Ciocalteau reagent dilute with 50 ml with DW
Highly SensitiveReagents are stableMore concentratedCan be used for both Normal & Micro assay
• First protein is reacted with alkaline cupric sulfate in the presence of tartarate for 10 minutes at room temperature. During this incubation, a tetradentate copper complex forms from four peptide bonds and one atom of copper.
• Second a i.e. Folin-phenol reagent is added, it reduced to give intense blue color.
• Reaction involves reduction of the Folin reagent and oxidation of aromatic residues.
• (Conversion of Cupric Cu+2 to Cuprous Cu+ ions, Read the blue colour developed, at 610 nm for colorimeter)
• Specificity: The Lowry method is best used with protein concentrations of 0.01-1.0 mg/mL
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