Fluoroprofile, a fluorescence-based assay for rapid and sensitive quantitation of proteins in solution.
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 microg/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.
Mackintosh
James A
JA
FLUOROtechnics Pty Ltd., Sydney, NSW, Australia.
Veal
Duncan A
DA
Karuso
Peter
P
D001578:Benzopyrans / Q000737:chemistry*; D005456:Fluorescent Dyes; D005663:Furans / Q000737:chemistry*; D007659:Ketones / Q000737:chemistry*; D011506:Proteins / Q000032:analysis*; D011933:Reagent Kits, Diagnostic; D012710:Serum Albumin, Bovine / Q000032:analysis; D013050:Spectrometry, Fluorescence / Q000379:methods
D001578:Benzopyrans; D005456:Fluorescent Dyes; D005663:Furans; D007659:Ketones; D011506:Proteins; D011933:Reagent Kits, Diagnostic; D012710:Serum Albumin, Bovine; C477464:epicocconone