title: Metal Enhanced Intrinsic Fluorescence of Proteins and Label-Free Bioassays.
abstract: Most of the applications of fluorescence require the use of labeled drugs and labeled biomolecules. Due to the need of labeling biomolecules with extrinsic fluorophores, there is a rapidly growing interest in methods which provide label-free detection (LFD). Proteins are highly fluorescent, which is due primarily to tryptophan residues. However, since most proteins contain tryptophan, this emission is not specific for proteins of interest in a biological sample. This is one of the reasons of not utilizing intrinsic tryptophan emission from proteins to detect specific proteins. Here, we present the intrinsic fluorescence for several proteins bound to the silver or aluminum metal nanostructured surfaces. We demonstrate the metal enhanced fluorescence (MEF) of proteins with different numbers of tryptophan residues. Large increases in fluorescence intensity and decreases in lifetime provide the means of direct detection of bound protein without separation from the unbound. We present specific detection of individual types of proteins and measure the binding kinetics of proteins such as IgG and streptavidin. Additionally, specific detection of IgG and streptavidin has been accomplished in the presence of large concentrations of other proteins in sample solutions. These results will allow design of surface-based assays with biorecognitive layer that specifically bind the protein of interest and thus enhance its intrinsic fluorescence. The present study demonstrates the occurrence of MEF in the UV region and thus opens new possibilities to study tryptophan-containing proteins without labeling with longer wavelength fluorophores and provides an approach to label-free detection of biomolecules.
authors.affiliation: Center for Fluorescence Spectroscopy, University of Maryland at Baltimore, Department of Biochemistry and Molecular Biology, 725 West Lombard Street, Baltimore, MD 21201, USA.
authors.forename: Krishanu
authors.initials: K
authors.lastname: Ray
authors.forename: Henryk
authors.initials: H
authors.lastname: Szmacinski
authors.forename: Mustafa H
authors.initials: MH
authors.lastname: Chowdhury
authors.forename: Joseph R
authors.initials: JR
authors.lastname: Lakowicz