Measuring plant protein with the Bradford assay : 1. Evaluation and standard method

The suitability of the Bradford protein assay for measuring plant protein was evaluated and a standard method developed. The assay involves extraction of dried, fresh, or frozen plant material in 0.1 NaOH for 30 min. Replicate 100-μl aliquots of centrifuged supernatant are assayed with 5 ml Bio-Rad Bradford dye reagent (Coomassie brilliant blue G-250) diluted 1:4 and containing 3 mg/ml soluble polyvinylpyrollidone. Absorbance at 595 nm is recorded after 15 min against an NaOH blank. Samples are calibrated against a ribulose 1,5-diphosphate carboxylase-oxygenase standard in NaOH. Procedures for plant preparation, extraction stability, the effects of phenol removal and quinone formation, and assay recovery are evaluated. Assay absorbance stability and techniques for increasing absorbance stability are reported. Changes in protein quality are briefly discussed.