Single-molecule spectroscopy for molecular identification in capillary electrophoresis.
The electrophoretic mobilities of individual DNA molecules were determined by using fluorescence imaging. The average mobility agreed well with that obtained in capillary electrophoresis (CE). The signal-to-noise ratio (S/N) did not decrease in the presence of up to 8% plasma or 8% raw blood. Single-molecule detection was still possible in the presence of 50% raw blood. Single-molecule CE of two differently labeled molecules was carried out in the presence of a transmission grating. Even when the mobility difference is not sufficient because of low S/N, identification using different fluorescence wavelengths can be performed at >99% accuracy. So, when small differences in DNA sequence due to disease or mutation can lead to hybridization to labels with different dyes, the screening of the mutated DNA will be facilitated by online spectroscopy in addition to the electrophoretic information from CE.
Lee
Ji-Young
JY
Ames Laboratory-US Department of Energy and Department of Chemistry, Iowa State University, Ames, IA 50011, USA.
Li
Hung-Wing
HW
Yeung
Edward S
ES
D019075:Electrophoresis, Capillary / Q000379:methods*; D013050:Spectrometry, Fluorescence