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

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.

Graphical abstractAn Aluminum micro-nanofluidic chip and a confocal optical detector were developed for sensitive real-time clinical pathogen molecular diagnostics. This is the first report of Al metal being used to build a lab-on-a-chip. The Al micro-nanofluidic chip had a surface contact angle of 93.8° and was inert to biomolecules. DNA amplification could be performed in a reaction volume as small as 39 nL, and a detection limit of approximately three genomic copies for stable and specific pathogen identification was obtained. Our results indicate that the chip is promising for applications in nanobiotechnology, nanomedicine, and clinical pathogen molecular diagnostics for nanoliter droplet amplification of several genomic copies from a single bacterium.Figure optionsDownload full-size imageDownload as PowerPoint slide