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Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique

One of the most important goals in proteomics is to detect the real-time kinetics of diverse biomolecular interactions. Fluorescence, which requires extrinsic tags, is a commonly and widely used method because of its high convenience and sensitivity. However, in order to maintain the conformational...

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Autores principales: Sun, Yung-Shin, Zhu, Xiangdong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2014
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4299064/
https://www.ncbi.nlm.nih.gov/pubmed/25490591
http://dx.doi.org/10.3390/s141223307
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author Sun, Yung-Shin
Zhu, Xiangdong
author_facet Sun, Yung-Shin
Zhu, Xiangdong
author_sort Sun, Yung-Shin
collection PubMed
description One of the most important goals in proteomics is to detect the real-time kinetics of diverse biomolecular interactions. Fluorescence, which requires extrinsic tags, is a commonly and widely used method because of its high convenience and sensitivity. However, in order to maintain the conformational and functional integrality of biomolecules, label-free detection methods are highly under demand. We have developed the oblique-incidence reflectivity difference (OI-RD) technique for label-free, kinetic measurements of protein-biomolecule interactions. Incorporating the total internal refection geometry into the OI-RD technique, we are able to detect as low as 0.1% of a protein monolayer, and this sensitivity is comparable with other label-free techniques such as surface plasmon resonance (SPR). The unique advantage of OI-RD over SPR is no need for dielectric layers. Moreover, using a photodiode array as the detector enables multi-channel detection and also eliminates the over-time signal drift. In this paper, we demonstrate the applicability and feasibility of the OI-RD technique by measuring the kinetics of protein-protein and protein-small molecule interactions in sandwich assays.
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spelling pubmed-42990642015-01-26 Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique Sun, Yung-Shin Zhu, Xiangdong Sensors (Basel) Article One of the most important goals in proteomics is to detect the real-time kinetics of diverse biomolecular interactions. Fluorescence, which requires extrinsic tags, is a commonly and widely used method because of its high convenience and sensitivity. However, in order to maintain the conformational and functional integrality of biomolecules, label-free detection methods are highly under demand. We have developed the oblique-incidence reflectivity difference (OI-RD) technique for label-free, kinetic measurements of protein-biomolecule interactions. Incorporating the total internal refection geometry into the OI-RD technique, we are able to detect as low as 0.1% of a protein monolayer, and this sensitivity is comparable with other label-free techniques such as surface plasmon resonance (SPR). The unique advantage of OI-RD over SPR is no need for dielectric layers. Moreover, using a photodiode array as the detector enables multi-channel detection and also eliminates the over-time signal drift. In this paper, we demonstrate the applicability and feasibility of the OI-RD technique by measuring the kinetics of protein-protein and protein-small molecule interactions in sandwich assays. MDPI 2014-12-05 /pmc/articles/PMC4299064/ /pubmed/25490591 http://dx.doi.org/10.3390/s141223307 Text en © 2014 by the authors; licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Sun, Yung-Shin
Zhu, Xiangdong
Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique
title Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique
title_full Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique
title_fullStr Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique
title_full_unstemmed Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique
title_short Real-Time, Label-Free Detection of Biomolecular Interactions in Sandwich Assays by the Oblique-Incidence Reflectivity Difference Technique
title_sort real-time, label-free detection of biomolecular interactions in sandwich assays by the oblique-incidence reflectivity difference technique
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4299064/
https://www.ncbi.nlm.nih.gov/pubmed/25490591
http://dx.doi.org/10.3390/s141223307
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