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Detection of Viable Pathogens Using Label-Free Electrical Detection of Nucleic Acid Amplification
Detection of Viable Pathogens Using Label-Free Electrical Detection of Nucleic Acid Amplification
Leveraging developments on electrical label-free detection of amplification and expertise in microfluidic systems, we plan to continue the development of an automated microchip for detection of pathogens. Our group has previously demonstrated specific detection of food borne pathogens in a silicon microarray. A primer dehydration protocol enables detection of multiple genes when the sample is partitioned in micro-reactors with different primer sets. In addition, we have shown that the amplification reaction can be electronically detected using ion-sensitive field effect transistors that monitor the electrolyte acidity. We now look to take advantage of the intrinsic scalability of electronics and the arraying technologies that we have developed to do electrical detection of multiple reactions. This will require creating platforms for parallel measurement of transistors and the development of a robust on-chip reference electrode that overcomes the current requirement of having an off-chip electrode which is limiting the number of parallel reactions that can be performed. Additionally, the incorporation of a new pre-amplification step will discriminate live vs. dead pathogens. We will study on-chip performance of viable qLAMP and the viability of performing the treatment on a microfluidic device. In this way, using electronics and microfluidics, we will develop a DNA amplification system for the detection of foodborne pathogens outside a laboratory setup.
Fig. Flow chart of the proposed approached for electrical detection of qLAMP. i) Starting from a sample of either grown or concentrated pathogens. ii) The PMA treatment is applied to inhibit amplification of DNA templates coming from compromised bacteria. iii) The treated sample is the partitioned in micro-droplets. Each pixel has a different set of primers for multiplexed detection. iv) The chip is heated to the reaction temperature and, v) An electrical signal is recorded before and after reactions to assess amplification based on surface potential changes.
