A microfluidic biosensor based on nucleic acid sequence recognition

被引:71
作者
Kwakye, S [1 ]
Baeumner, A [1 ]
机构
[1] Cornell Univ, Dept Biol & Environm Engn, Ithaca, NY 14853 USA
基金
美国国家科学基金会;
关键词
biosensor; microfluidics; rapid detection; probe hybridization; microfabrication; liposomes;
D O I
10.1007/s00216-003-2063-2
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
The development of a generic semi-disposable microfluidic biosensor for the highly sensitive detection of pathogens via their nucleic acid sequences is presented in this paper. Disposable microchannels with defined areas for capture and detection of target pathogen RNA sequence were created in polydimethylsiloxane (PDMS) and mounted onto a reusable polymethylmethacrylate (PMMA) stand. Two different DNA probes complementary to unique sequences on the target pathogen RNA serve as the biorecognition elements. For signal generation and amplification, one probe is coupled to dye encapsulated liposomes while the second probe is coupled to superparamagnetic beads for target immobilization. The probes hybridize to target RNA and the liposome-target-bead complex is subsequently captured on a magnet. The amount of liposomes captured correlates directly to the concentration of target sequence and is quantified using a fluorescence microscope. Dengue fever virus serotype 3 sequences and probes were used as a model analyte system to test the sensor. Probe binding and target capture conditions were optimized for sensitivity resulting in a detection limit of as little as 10 amol muL(-1) (10 pmol L-1). Future biosensors will be designed to incorporate a mixer and substitute the, fluorescence detection with an electrochemical detection technique to provide a truly portable microbiosensor system.
引用
收藏
页码:1062 / 1068
页数:7
相关论文
共 23 条
[1]   RNA biosensor for the rapid detection of viable Escherichia coli in drinking water [J].
Baeumner, AJ ;
Cohen, RN ;
Miksic, V ;
Min, JH .
BIOSENSORS & BIOELECTRONICS, 2003, 18 (04) :405-413
[2]   Biosensor for Dengue virus detection: Sensitive, rapid, and serotype specific [J].
Baeumner, AJ ;
Schlesinger, NA ;
Slutzki, NS ;
Romano, J ;
Lee, EM ;
Montagna, RA .
ANALYTICAL CHEMISTRY, 2002, 74 (06) :1442-1448
[3]   Nanoparticles with Raman spectroscopic fingerprints for DNA and RNA detection [J].
Cao, YWC ;
Jin, RC ;
Mirkin, CA .
SCIENCE, 2002, 297 (5586) :1536-1540
[4]   Development of a laser-induced cell lysis system [J].
Dhawan, MD ;
Wise, F ;
Baeumner, AJ .
ANALYTICAL AND BIOANALYTICAL CHEMISTRY, 2002, 374 (03) :421-426
[5]   Rapid prototyping of microfluidic systems in poly(dimethylsiloxane) [J].
Duffy, DC ;
McDonald, JC ;
Schueller, OJA ;
Whitesides, GM .
ANALYTICAL CHEMISTRY, 1998, 70 (23) :4974-4984
[6]  
*DYN INC, 1998, BIOM TECHN MOL BIOL
[7]   The BARC biosensor applied to the detection of biological warfare agents [J].
Edelstein, RL ;
Tamanaha, CR ;
Sheehan, PE ;
Miller, MM ;
Baselt, DR ;
Whitman, LJ ;
Colton, RJ .
BIOSENSORS & BIOELECTRONICS, 2000, 14 (10-11) :805-813
[8]   Detection of viable Cryptosporidium parvum using DNA-modified liposomes in a microfluidic chip [J].
Esch, MB ;
Locascio, LE ;
Tarlov, MJ ;
Durst, RA .
ANALYTICAL CHEMISTRY, 2001, 73 (13) :2952-2958
[9]   Detection of Cryptosporidium parvum using oligonucleotide-tagged liposomes in a competitive assay format [J].
Esch, MB ;
Baeumner, AJ ;
Durst, RA .
ANALYTICAL CHEMISTRY, 2001, 73 (13) :3162-3167
[10]   Biosensor for the specific detection of a single viable B-anthracis spore [J].
Hartley, HA ;
Baeumner, AJ .
ANALYTICAL AND BIOANALYTICAL CHEMISTRY, 2003, 376 (03) :319-327