Very Quick Reverse Transcription Polymerase Chain Reaction for Detecting 2009 H1N1 Influenza A Using Wire-Guide Droplet Manipulations

This technology comprises a reprogrammable digital microfluidics assembly, called wire-guided droplet microfluidics, wherein a droplet hangs at the end of a wire or a blunt end syringe needle and moves either over a superhydrophobic surface or within a hydrophobic liquid. The differences in the forces of adhesion to the needle wire and to the superhydrophobic surface/liquid enables the movements of a droplet following the movements of the needle wire. The wire-guided droplet microfluidics, is totally reprogrammable and simple enough to be fabricated into a handheld device that can be used in field situations. The whole processes of PCR assay can be performed in a single device without using any human labor. The entire analysis can be finished in less than 15 min including serial dilution, centrifuging, DNA extraction, thermocycling and real-time quantification, all with wire-guided droplet microfluidics in a single device.
 

Background

There have been many attempts of incorporating the complete process of polymerase chain reaction (PCR) assays into microchannels. The problem of these attempts is that the user cannot change the assay protocol or chip architecture easily.
 

Advantages

• Simplicity
  • Complicated fabrication (e.g. electrowetting) of a microfluidic device is not necessary
  • No magnetic particles (e.g. magnetofluidics) are needed that may affect the assays
• Low-cost - does not involve high-end equipment
  • Uses mostly open-source electronics, such as Arduino 
  • The material/equipment cost to build the prototype device is estimated as low as <$250
• Demonstration of dilution, centrifuging, DNA extraction and real-time quantification in addition to thermocycling
  • All other known droplet microfluidic PCR assays are focused on thermocycling. This invention includes serial dilution, centrifuging, DNA extraction and real-time quantification, in addition to very rapid thermocycling
• Reprogrammability
  • The proposed device demonstrates the adaptability of the system to replace many different types of common laboratory tasks on a single platform, and even interrupt the programmed protocol and implement an additional process in rapid succession
• Sensitivity
  • Unlike the other microfluidic PCR approaches that uses sub-μL or nL volume, this method uses 1 or 10 μL volume, which does not sacrifice the assay sensitivity. With 10 nL volume, for example, the lowest detectable concentration is 100,000 cells/mL, while with 10 μL volume, it is 100 cells/mL.

Applications

• Can be used for a wide variety of common laboratory tasks
• Can be used to facilitate all steps of PCR assays, including serial dilution, centrifugation, gene extraction, thermocycling and its real-time quantification, enabling fully automated and reprogrammable PCR assays
• Can be used to detect COVID-19

Status: issued U.S. patent #9,719,134

COVID, COVID-19, COVID19, Coronavirus