Integrated Microfluidic Electrophoresis System for Analysis of Genetic Materials Using Signal Amplification Methods

• Published in: Analytical chemistry (Washington) 2002-02, Vol.74 (4), p.725-733
• Main Authors: Tang, Thompson, Badal, M. Youssouf, Ocvirk, Gregor, Lee, William E, Bader, Douglas E, Bekkaoui, Faouzi, Harrison, D. Jed
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Cell physiology; Chemistry; Deoxyribonucleic acid; DNA; DNA - analysis; DNA Probes - analysis; DNA, Bacterial - analysis; Electrophoresis; Fundamental and applied biological sciences. Psychology; Indicators and Reagents; Methicillin Resistance; Microcomputers; Molecular and cellular biology; Nanotechnology; Reverse Transcriptase Polymerase Chain Reaction; Signal transduction; Staphylococcus aureus - genetics
• ISSN: 0003-2700; 1520-6882
• DOI: 10.1021/ac010874j

An isothermal signal amplification technique for specific DNA sequences, known as cycling probe technology (CPT), was performed within a microfluidic chip. The presence of DNA from methicillin-resistant Staphylococcus aureus was determined by signal amplification of a specific DNA sequence. The microfluidic device consisted of four channels intersecting to mix the sample and reagents within 55 s, as they were directed toward the reactor coil by electrokinetic pumping. The 160-nL CPT reactor occupied ∼220 mm2. Gel-free capillary electrophoresis separation of the biotin- and fluorescein-labeled probe from the probe fragments was performed on-chip following the on-chip reaction. An off-chip CPT reaction, with on-chip separation gave a detection limit of 2 fM (0.03 amol) target DNA and an amplification factor of 85 000. Calibration curves, linear at