Systems and methods for continuous flow digital droplet polymerase chain reaction bioanalysis

Abstract

Systems and methods for continuous flow polymerase chain reaction (PCR) are provided. The system comprises an injector, a mixer, a coalescer, a droplet generator, a detector, a digital PCR system, and a controller. The injector takes in a sample, partitions the sample into sample aliquots with the help of an immiscible oil phase, dispenses waste, and sends the sample aliquot to the mixer. The mixer mixes the sample aliquot with a PCR master mix and diluting water, dispenses waste, and sends the sample mixture (separated by an immiscible oil) to the coalescer. The coalescer coalesces the sample mixture with primers dispensed from a cassette, dispenses waste, and sends the reaction mixture (separated by an immiscible oil) to the droplet generator. The droplet generator converts the sample mixture into an emulsion where aqueous droplets of the reaction mixture are maintained inside of an immiscible oil phase and dispenses droplets to the digital PCR system. The digital PCR system amplifies target DNAs in the droplets. The detector detects target DNAs in the droplets. The controller controls the system to run automatically and continuously.