INTERFACIAL CONSIDERATIONS FOR DROPLET PCR LAB-ON-CHIP DEVICES

Abstract

Lab-on-chip devices have the potential to decentralize the current model of diagnostics to point-of-care diagnostics. Easy to use, low cost, rapid infectious disease diagnostic tools could especially impact and improve healthcare in low resource areas. Micro-total-analytical-systems could also enable smarter medical decisions, quicker patient recoveries, and cheaper healthcare costs in fully developed settings. Significant innovations to standard technologies used today will help realize the promise of lab-on-chip devices. In this work, innovative technologies compatible with current lab-on-chip devices were investigated to simplify their operation, decrease their complexity, and reduce their cost. The interfacial aspects that dominate microfluidic systems, and in particular droplet polymerase chain reaction (PCR) devices, are emphasized.

Droplet PCR utilizing microfluidic technology has largely been automated, but sample preparation methods prior to amplification remains a laborious process. We have developed particles that condense the many steps of sample preparation into a single buffer protocol. The particles were made by crosslinking chitosan, a pH responsive biopolymer. DNA was electrostatically and sterically adsorbed to the beads at pH 8.5. Furthermore, amplification of DNA directly off the beads was demonstrated eliminating the need to desorb DNA into solution. Implementation of these particles will drastically simplify droplet PCR lab-on-chip devices.

We also characterized the adsorption of polymerase at the oil-water interfaces of droplets and identified a surfactant to prevent the loss of polymerase in solution. The pendant drop technique was used to observe the change in interfacial tension due to adsorption of Taq Pol and/or surfactants to the interface. PCR performance of two surfactants, Brij L4 and ABIL EM90, were predicted from equilibrium interfacial tension measurements. Brij L4, a surfactant that had never been used with PCR, prevented polymerase adsorption and enabled more efficient PCR than ABIL EM90, a popular PCR surfactant.

Lastly, we ambitiously designed a system to conduct droplet PCR without oil or surfactants. Droplets were generated on-chip by adapting a co-flow droplet generating device previously developed in our group. Then droplets were immobilized on-chip in hydrodynamic traps. Two different modes of trapping were demonstrated, indirect and direct. Also, all aspects of an air continuous phase droplet PCR device were considered such as protein adsorption to channel walls and droplet evaporation during thermal cycling.