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bioRxiv · 10.64898/2026.03.03.709423

A solid-state heater-imager for quantitative evaluation of colorimetric isothermal nucleic acid amplification on paper

Raut, B.·Palla, G.·Nugyen, D. V.·Munds, R. A.·Bayram, A.·Kumar, V.·Ahmed, B.·Ault, A.·Gilbertie, A.·Pasternak, J. A.·Verma, M. S.
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Abstract

Maintaining precise isothermal conditions in portable nucleic acid amplification tests (NAATs) is critical for reproducible results but remains challenging with conventional single-sided thin-film heaters, which exhibit temperature gradients and strong dependence on ambient conditions. To close this gap, we engineered ThermiQuant VitroMini, a dual-sided heater design that achieves volumetric-level temperature uniformity using thin-film heaters while preserving optical transparency for real-time colorimetric loop-mediated isothermal amplification (LAMP) analysis on microfluidic paper-based analytical devices ({micro}PADs). The device integrates two independently regulated indium tin oxide (ITO) heaters (8 {Omega} each) controlled by independent proportional-integral-derivative (PID) algorithms. Heaters were evaluated under controlled ambient environments of 4 {degrees}C (refrigerated), 23 {degrees}C (room temperature), and 50 {degrees}C (oven). Analytical tests were performed using a colorimetric LAMP assay targeting the SARS-CoV-2 orf7ab gene on {micro}PADs preloaded with dried LAMP reagents, with time-lapse images (30 seconds interval) analyzed via Amplimetrics software. VitroMini maintained 65 {+/-} 0.5 {degrees}C across 4 to 50 {degrees}C ambient conditions and achieved a limit of detection of 34 copies/reaction (4.5 copies/{micro}L) and limit of quantification of 1000 copies/reaction (133 copies/{micro}L), with quantification time (Tq) linearly correlated with log10 DNA concentration. Dual-sided heating eliminated temperature bias, condensation artifacts, and ambient-dependent variability while preserving optical transparency for real-time quantitative LAMP reaction. ThermiQuant VitroMini bridges the gap between benchtop volumetric heaters and portable diagnostic devices, offering a compact, low-power platform for quantitative colorimetric molecular analysis on paper with potential for decentralized and field-deployable applications.

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Raut, B., Palla, G., Nugyen, D. V., Munds, R. A., Bayram, A., Kumar, V., Ahmed, B., Ault, A., Gilbertie, A., Pasternak, J. A., Verma, M. S.. 2026-03-09. A solid-state heater-imager for quantitative evaluation of colorimetric isothermal nucleic acid amplification on paper. https://doi.org/10.64898/2026.03.03.709423

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