Quantitative Analysis of Nucleic Acid Hybridization on Magnetic Particles and Quantum Dot-Based Probes 英文参考文献.docVIP

Sensors 2009, 9, 5590-5599; doi:10.3390/OPEN ACCESS sensors ISSN 1424-8220 /journal/sensors Article Quantitative Analysis of Nucleic Acid Hybridization on Magnetic Particles and Quantum Dot-Based Probes Sun Hee Lim 1,*, Felix Bestvater 2, Philippe Buchy 3, Sek Mardy 3, and Alexey Dan Chin Yu 1,* 1 Diagnostics, Institut Pasteur Korea, 696 Sampyeong-dong, Bundang-gu, Seongnam City, Gyeonggi- do, Korea, 443-270 2 Cell Biophysics, Institut Pasteur Korea, 696 Sampyeong-dong, Bundang-gu, Seongnam City, Gyeonggi-do, Korea, 443-270; E-Mail: f.bestvater@ 3 Virology Unit, Institut Pasteur in Cambodia, 5, Monivong boulevard, Phnom Penh, Cambodia; E-Mails: pbuchy@ (P.B.); mardy@ (S.M.) * Authors to whom correspondence should be addressed; E-Mails: sunhlim@ (S.H.L.); alexey.dan@ (A.D.C.Y.); Tel.: +82-8018-8300; Fax: +82-31-8018-8013 Received: 17 June 2009; in revised form: 8 July 2009 / Accepted: 13 July 2009 / Published: 14 July 2009 Abstract: In the present study we describe sandwich design hybridization probes consisting of magnetic particles (MP) and quantum dots (QD) with target DNA, and their application in the detection of avian influenza virus (H5N1) sequences. Hybridization of 25-, 40-, and 100-mer target DNA with both probes was analyzed and quantified by flow cytometry and fluorescence microscopy on the scale of single particles. The following steps were used in the assay: (i) target selection by MP probes and (ii) target detection by QD probes. Hybridization efficiency between MP conjugated probes and target DNA hybrids was controlled by a fluorescent dye specific for nucleic acids. Fluorescence was detected by flow cytometry to distinguish differences in oligo sequences as short as 25-mer capturing in target DNA and by gel-electrophoresis in the case of QD probes. This report shows that effective manipulation and control of micro- and nanoparticles in hybridization assays is