Journal Article

ThermiQuant™ AquaStream: A portable instrument for quantitative colorimetric isothermal nucleic acid amplification reactions in paper and tube formats

Kumar, Virendra, Wang, Jiangshan, Campbell, Skyler, Ra, Kyungyeon, Raut, Bibek, Palla, Gopal, Verma, Mohit S.

2026

Overview

Microfluidic paper-based analytical devices (µPADs) are an attractive format for colorimetric nucleic acid amplification tests (NAATs) because they enable low-cost, portable diagnostics in resource-limited settings. However, researchers often optimize colorimetric assays in liquid reactions in tubes before translating them to µPADs. Since both formats require separate instruments for incubation and real-time sensing, direct comparison of reactions between the two formats is difficult. To address these cross-platform limitations, we developed ThermiQuant™ AquaStream, a portable benchtop device (15 × 20 × 16 cm, ~ 5 kg; cost: USD 327) that supports seamless colorimetric loop-mediated isothermal amplification (LAMP) reactions in both µPADs and tubes under a common workflow. The system enables real-time reaction tracking (every 30 seconds) through onboard image processing, precise isothermal control (±0.5 °C) using a repurposed consumer-grade sous-vide heater, and medium-throughput (24 tubes or 42 µPADs). Testing with synthetic SARS-CoV-2 orf7ab DNA fragments demonstrated a limit of detection corresponding to a 95% probability of detection (LOD95) of 110 copies per reaction in tube (22 copies/µL) and 39 copies per reaction in µPADs (5 copies/µL), estimated using probit regression. In both formats the limit of quantification (LOQ), defined as the lowest concentration yielding a coefficient of variation (CV) of quantification time (Tq) ≤ 10%, was 250 copies/reaction resulting in a strong linear (R 2 = 0.98 & R 2 = 0.96 respectively for tube and µPADs) standard calibration curves. ThermiQuant™ AquaStream provides an affordable and versatile benchtop platform capable of supporting both tube- and µPAD-based colorimetric LAMP assays, serving as a proof-of-concept research tool for assay development and molecular diagnostics in One Health settings such as clinics, farms, and field environments.

• Publisher: Public Library of Science,PLOS,Public Library of Science (PLoS)
• Subject: Assaying / Automation / Biology and Life Sciences / Cameras / Coefficient of variation / Colorimetric analysis / Colorimetry / Colorimetry - instrumentation / Colorimetry - methods / Comparative analysis / Computer and Information Sciences / Design / Dyes / Engineering and Technology / Gene amplification / Humans / Image processing / Medicine and health sciences / Methods / Microfluidics / Molecular Diagnostic Techniques - instrumentation / Molecular Diagnostic Techniques - methods / Nucleic Acid Amplification Techniques - instrumentation / Nucleic Acid Amplification Techniques - methods / Nucleic acids / Physical Sciences / Portable equipment / Rapid Diagnostic Tests / Real time / Research and Analysis Methods / SARS-CoV-2 - genetics / SARS-CoV-2 - isolation & purification / Severe acute respiratory syndrome coronavirus 2 / Smartphones / Statistical analysis / Thermodynamic processes / Tubes / Water heaters