Unit

UNIT 10.1A Spectrophotometric and Colorimetric Determination of Protein Concentration

  1. Michael H. Simonian (spectrophotometric methods)1,
  2. John A. Smith (colorimetric methods)2

Published Online: 1 MAY 2001

DOI: 10.1002/0471142727.mb1001as35

Current Protocols in Molecular Biology

Current Protocols in Molecular Biology

How to Cite

Simonian, M. H. and Smith, J. A. 2001. Spectrophotometric and Colorimetric Determination of Protein Concentration. Current Protocols in Molecular Biology. 10:I:10.1A.

Author Information

  1. 1

    Beckman Coulter, Inc., Fullerton, California

  2. 2

    University of Alabama at Birmingham, Birmingham, Alabama

Publication History

  1. Published Online: 1 MAY 2001
  2. Published Print: JUL 1996

This is not the most recent version of the article. View current version (1 NOV 2006)

Abstract

This unit describes spectrophotometric and colorimetric methods for measuring the concentration of a sample protein in solution. Absorbance measurement at 280 nm is used to calculate protein concentration by comparison with a standard curve or published absorptivity values for that protein. An alternate protocol uses absorbance at 205 nm to calculate the protein concentration. Both methods can be used to quantitate total protein in crude lysates and purified or partially purified protein. Use of a spectrofluorometer or a filter fluorometer to measure the intrinsic fluorescence emission of a sample solution is also described. The measurement is compared with the emissions from standard solutions to determine the concentration of purified protein. The Bradford colorimetric method, based upon binding of the dye Coomassie brilliant blue to an unknown protein, is also presented, as is the Lowry method, which measures colorimetric reaction of tyrosyl residues in an unknown.