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Proceedings Paper

Quantitative fluorescence resonance energy transfer measurements using microarray technology
Author(s): Jiang Zhu; Cheng Deng; Guoliang Huang; Yang Yang; Shukuan Xu; Zhonghua Dong; Xiaoyong Yang; Keith Mitchelson; Jing Cheng
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Paper Abstract

Fluorescence resonance energy transfer (FRET) has found wide use in structural biology, biochemistry, and cell biology for measuring intra- and inter-molecular distances in the 1-10 nm range and for obtaining quantitative spatial and temporal information about the interaction of proteins, lipids, and DNA. The measurements of distances and interactions are based on the calculation of the fluorescence transfer efficiency using some algorithms to process the acquired images from several different filter sets. However, FRET measurements can suffer from several sources of distortion because of cross talk between donor and acceptor fluorophores. In this paper, we measured the FRET efficiency on glass coverslips using microarray technology and described an algorithm to analyze the FRET data obtained, which is corrected for the cross talk due to spectral overlap of donor and acceptor molecules. Measurement of the interaction of the donor and acceptor, which are mixed together or coupled to the respective 3'-end and 5'-end of a single-strand DNA are shown to document the accuracy of the approach, and allow one to estimate cross talk between the different filter units and to reveal the relationship of the FRET efficiencies of these two samples relative to the donor and acceptor concentrations.

Paper Details

Date Published: 1 May 2007
PDF: 7 pages
Proc. SPIE 6534, Fifth International Conference on Photonics and Imaging in Biology and Medicine, 653419 (1 May 2007); doi: 10.1117/12.741482
Show Author Affiliations
Jiang Zhu, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Cheng Deng, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Guoliang Huang, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Yang Yang, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Shukuan Xu, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Zhonghua Dong, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Xiaoyong Yang, National Engineering Research Ctr. for Beijing Biochip Technology (China)
Keith Mitchelson, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)
Jing Cheng, Tsinghua Univ. School of Medicine (China)
National Engineering Research Ctr. for Beijing Biochip Technology (China)


Published in SPIE Proceedings Vol. 6534:
Fifth International Conference on Photonics and Imaging in Biology and Medicine

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