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Ultra-fast, sensitive and quantitative on-chip detection of group B streptococci in clinical samples
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| Abstract |
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| PCR enables sensitive and specific detection of infectious disease agents, but application in point-of-care diagnostic testing remains scarce. A compact tool that runs PCR assays in less than a few minutes and that relies on mass-producible, disposable reactors could revolutionize while-you-wait molecular testing. We here exploit well-established semiconductor manufacturing processes to produce silicon ultra-fast quantitative PCR (UF-qPCR) chips that can run PCR protocols with limited assay optimization. A total of 110 clinical samples were analyzed for the detection of group B streptococci using both a validated benchtop and an on-chip qPCR assay. For the onchip assay, the total reaction time was reduced after optimization to less than 5 min. The standard curve, spanning a concentration range of S log units, yielded a PCR efficiency of 94%. The sensitivity obtained was 96% (96/100; CI: 90-98%) and the specificity 70% (7/10; CI: 40-90%). We show that if melting analyses would be integrated, the obtained sensitivity would drop slightly to 93% (CI: 86-96%), while the specificity would increase to 100% (CI: 72% - 100%). In comparison to the benchtop reference qPCR assay performed on a LightCycler(C)96, the on-chip assay demonstrated a highly significant qualitative (Spearman's rank correlation) and quantitative (linear regression) correlation. Using a mass-producible qPCR chip and limited assay optimization, we were able to develop a validated qPCR protocol that can be carried out in less than five minutes. The analytical performance of the microchip-based UF-qPCR system was shown to match that of a benchtop assay. This is the first report to provide UF-qPCR validation using clinical samples. We demonstrate that qPCR-based while-you-wait testing is feasible without jeopardizing assay performance. |
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English
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Talanta : the international journal of pure and applied analytical chemistry. - Oxford, 1958, currens | |
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Oxford : Pergamon, 2019
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0039-9140 [print]
1873-3573 [online]
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192(2019), p. 220-225
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000449443900030
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30348381
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