Sensitive sequence-specific molecular identification system comprising an aluminum micro-nanofluidic chip and associated real-time confocal detector

摘要

We developed a micro-nanofluidic bioreactor-detector system for isothermal DNA amplification and sensitive real-time detection of the amplified products for sequence-specific molecular identification. Aluminum (Al) chips with a range of volumes from 7.07 μL to 39 nL and an associated real-time confocal optical detector are described. The detector provided highly sensitive fluorescence detection and low background noise. One of the important aspects of the system was the development of a surface processing technique that afforded chips with an inert surface to improve amplification stability in micro-nanoliter reaction assays. The micro-nanofluidic system exhibited more sensitive exponential DNA amplification characteristics than a standard PCR tube amplification system with a volume of 25 μL, the response time was clearly reduced at the same DNA template concentration, and the sensitivity in the number of copies of the DNA template was improved by >600-fold. Efficient amplification of nucleic acid was achieved with as few as three copies of the DNA template. This system may be useful for the development of novel lab-on-a-chip devices and shows promise for single-molecule amplification in droplet assays, with potential applications in nanobiotechnology, nanomedicine, and clinical molecular diagnostics.