Self-Digitization Microfluidic Platform for Digital PCR

Project Summary AbstractThis SBIR Direct Phase II project will produce a complete platform to perform nucleic acid quantificationNAQby digital polymerase chain reactiondPCRThe platform will consist of a complete instrument that handles devices and executes all phases of the assaydigitizationamplificationand detectionin one instrumentoffering truewalk awaycapabilities within a closed systemIt will also include a consumable chip specifically tailored to high sensitivity applications such as circulating tumor DNActDNAresearch and diagnosticsa rapidly growing market projected to exceed $billion byThe platform will be valuable for biomedical research and clinical diagnostic applicationswith specific health related impact in areas such as cancer diagnostics and treatment monitoringinfectious disease diagnosticsviral load monitoring and moreThe platform is based on Self DigitizationSDtechnologya simplerobust method that efficiently uses a network of channels and wells to spontaneously partition samples into an array of predefined compartmentsThis platform is ideally suited for digital polymerase chain reactiondPCRapplicationsdPCR works by partitioning samples into hundredsthousandsor even millions of individual volumes where each volume may or may not contain target DNAOnly volumes with target DNA give a positive signal resulting in adigitalyes no signalfrom which Poisson statistics can then give direct determination of sample concentrationsDigital PCR directly provides absolute quantificationis robust against variations in reaction efficiencyis incredibly sensitiveand has nearly unlimited resolution capabilitiesmaking it technically superior to the current NAQgold standardof qPCRHoweverno commercial dPCR system is currently able to be compete with qPCR because of issues with workflowthroughputand costAmong dPCR platformsthe SD mechanism is uniquely suited to efficiently handle large sample volumeswhich provides a strong value proposition in highsensitivity applicationsand allows for direct competition with qPCR based assaysproviding a path to compete in both the dPCR and qPCR marketThe primary objective of this proposal is to produce a platform that overcomes the workflowthroughputcostand flexibility hurdles that have limited existing commercial dPCR systemsparticularly in their inability to penetrate the largerreal timeor qPCR marketThe first Aim will be development and construction of the instrumentincluding a digitization mechanism that will preparesamples in parallela high speed thermalcycling systeman imaging system to rapidly analyze samplesand integrated software to enable all stages from a singlegocommandThe second Aim will be development of materials and device designs to enable manufacturing of commercially viable devicesThe third Aim will encompass validation of the platform with clinically relevant and valuable NAQ assaysSpecificallyit will be validated with an assay relevant to ctDNA research and diagnostics Project Narrative This research will generate a digital PCRdPCRplatform to perform nucleic acid quantificationNAQconsisting of a stand alone instrument withwalk awaycapabilitiesand commercially viable devices to perform high sensitivity assaysThis instrument will offer unrivaled workflow capabilities and cost profiles that will transform the dPCR market and enable inroads into the larger qPCR marketIt will impact and enable many biological research and clinical applications including cancer diagnostics and treatment monitoringinfectious disease detectionand viral load monitoring