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Fluoroprofile, a fluorescence‐based assay for rapid and sensitive quantitation of proteins in solution
Author(s) -
Mackintosh James A.,
Veal Duncan A.,
Karuso Peter
Publication year - 2005
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200500095
Subject(s) - fluorescence , bradford protein assay , chemistry , bicinchoninic acid assay , chromatography , quantitative proteomics , biochemistry , proteomics , physics , quantum mechanics , gene
The development of a sensitive fluorescence‐based assay for the quantitative determination of protein concentration is described. The assay is based on the natural product epicocconone, which produces a large increase in fluorescence quantum yield upon binding to detergent‐coated proteins in solution. There is a concomitant shift in the emission maximum from 520 to 605 nm after binding, which results in low background signal allowing a linear dynamic range of 40 ng/mL to 200 μg/mL for most proteins. There is little protein‐to‐protein variation except for iron‐containing proteins and the assay can be used so that it is tolerant of chemicals commonly used in 2‐D sample buffers. The assay is more sensitive than standard absorption assays such as the Bradford and Lowry assays, and has a greater dynamic range and sensitivity than other fluorescent assays.