Sensing of p53 and EGFR Biomarkers Using High Efficiency SERS Substrates.

Peter Owens,Malini Olivo,Jayakumar Perumal,Gerard M O'Connor,Nigel Phillipson

doi:10.3390/bios5040664

Peter Owens, Malini Olivo + Show 3 more

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https://doi.org/10.3390/bios5040664

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Abstract

In this paper we describe a method for the determination of protein concentration using Surface Enhanced Raman Resonance Scattering (SERRS) immunoassays. We use two different Raman active linkers, 4-aminothiophenol and 6-mercaptopurine, to bind to a high sensitivity SERS substrate and investigate the influence of varying concentrations of p53 and EGFR on the Raman spectra. Perturbations in the spectra are due to the influence of protein–antibody binding on Raman linker molecules and are attributed to small changes in localised mechanical stress, which are enhanced by SERRS. These influences are greatest for peaks due to the C-S functional group and the Full Width Half Maximum (FWHM) was found to be inversely proportional to protein concentration.

Highlights

The development of ultrasensitive and rapid approaches to detect protein biomarkers at very low concentrations in a physiological environment represents a challenge in nano-medicine
Based on the very large improvement in signal intensity, silver Silmeco substrates were chosen for development of the Surface Enhanced Raman Resonance Scattering (SERRS) based immunoassay for p53 and Epidermal Growth Factor Receptor (EGFR)
We have shown that the influence of protein on the properties of specific peaks in Raman spectra can be measured and used as an evaluation of protein concentration

Summary

Introduction

The development of ultrasensitive and rapid approaches to detect protein biomarkers at very low concentrations in a physiological environment represents a challenge in nano-medicine. The most widely used method for analysis of low levels of protein in basic research and clinical diagnostics is the Enzyme-Linked Immunosorbent Assay (ELISA). In terms of diagnostic protein assays, Raman spectroscopic techniques with high sensitivity and multiplexing capabilities may offer competition to fluorescence based ELISA methods. Using Surfaced Enhanced Raman Spectroscopy (SERS) methods can in some cases surpass the measurement sensitivity of fluorescence. Intrinsic Raman spectra are inherently weak, the scattering cross-section can be greatly enhanced by positioning of the molecular species of interest near a nanostructured metal surface (typically gold or silver) [2,3]. Measurement sensitivities of up to 10−13 M have been reported and more flexible multiplexing protocols have been developed due to the very narrow spectral width of Raman peaks (typically 10–100 times narrower than fluorescence peaks) [5]

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