Quantitative fluorescence correction incorporating Förster resonance energy transfer and its use for measurement of hybridization efficiency on microarrays.
Fluorescence detection using two spectrally distinct fluorophores has long been used for the determination of the relative abundance of biomolecules, but overlap between the fluorescence spectra of each fluorophore can result in nonradiative Förster resonance energy transfer (FRET) and distorting the signals detected by fluorescence channels. Thus conventional methods for quantifying the relative abundance of fluorophores by fluorescence emission will not be accurate if FRET can occur. In this paper we report the development of a quantitative fluorescence correction method incorporating FRET to measure the relative abundance of fluorophores in dual-labeling experiments. The quantitative fluorescence correction method incorporating FRET is accurate, comprehensive, and convenient for the measurement of the relative abundance of fluorophores in dual-labeling experiments and can also correct the FRET distortion and provide accurate, quantitative, and convenient measurement of the hybridization efficiencies on microarrays.
Zhu
Jiang
J
Medical Systems Biology Research Center, Tsinghua University School of Medicine, Haidian District, Beijing 100084, PR China.
Deng
Cheng
C
Huang
Guoliang
G
Xu
Shukuan
S
Mitchelson
Keith
K
Cheng
Jing
J
D001483:Base Sequence; D003116:Color; D004247:DNA / Q000032:analysis / Q000737:chemistry / Q000235:genetics; D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer; D020869:Gene Expression Profiling; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013499:Surface Properties
D004247:DNA