Ultrafast plasmonic and real-time label-free polymerase chain reaction for point-of-care diagnostics

In response to the need of the world's medical community for accurate and immediate identification of infectious diseases, many researchers have focused on adapting the gold-standard molecular diagnostic method, polymerase chain reaction (PCR), to point-of-care (POC) applications.Despite all advances in microfluidic PCR as an alternative to bulky, slow, and powerintensive conventional thermocyclers, a simple-to-operate/fabricate PCR device is still lacking.In this thesis, we introduce a miniature plasmonic PCR thermocycler in which fast DNA amplification is derived from efficient photothermal heating of suspended gold nanorods (AuNRs) inside PCR reaction by a small-scale vertical-cavity surface-emitting laser (VCSEL).Using this method, we demonstrate 30 cycle-assay time of sub-ten-minute for successful Chlamydia Trachomatis DNA amplification in 20 L total PCR sample volume containing 2.5 nM AuNRs.Furthermore, we report an ultrasensitive real-time amplicon detection strategy which is based on cycle-by-cycle monitoring 260 nm absorption of PCR reaction.This was accomplished by irradiating PCR sample with a UV LED and collecting the transmitted optical power by a photodetector.UV absorption dependency on nucleotides' structural degree of freedom gives rise to distinctive features in the shape of UV amplification curves for PCR results determination (success/fail) and amplicon quantification with a high detection sensitivity of one DNA copy.This is the first demonstration of a compact plasmonic thermocycler combined with a real-time fluorophore-free quantitative amplicon detection system.The small footprint of the proposed PCR device stems from hardware miniaturization while abundant sample volume facilitates highly sensitive detection and fluid-handling required for on-field sample analysis.This simple, portable, and rapid PCR device is the first proof-of-concept demonstration of combining plasmonic DNA amplification and UV detection towards POC molecular diagnostics.More importantly, the realtime detection method is universal and purely optical solely dependent on standard PCR reagent (nucleotides) circumventing the need for the complicated and costly design of target-specific probes.

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