Low-Cost Arduino Reverse Transcriptase Loop-Mediated Isothermal Amplification (RT-LAMP) for Sensitive Nucleic Acid Detection.

This work presents a low-cost transcription loop-mediated isothermal amplification (RT-LAMP) instrument for nucleic acid detection, employing an Arduino Nano microcontroller. The cooling system includes customized printed circuit boards (PCBs) that serve as electrical resistors and incorporate fans. An aluminum block is designed to accommodate eight vials. The system also includes two PCB heaters-one for sample heating and the other for vial lid heating to prevent condensation. The color detection system comprises a TCS3200 color 8-sensor array coupled to one side of the aluminum heater body and a white 8-LED array coupled to the other side, controlled by two Multiplexer/Demultiplexer devices. LED light passes through the sample, reaching the color sensor and conveying color information crucial for detection. The top board is maintained at 110 ± 2 °C, while the bottom board is held at 65 ± 0.5 °C throughout the RT-LAMP assay. Validation tests successfully demonstrated the efficacy of the colorimetric RT-LAMP reactions using SARS-CoV-2 RNA amplification as a sample viability test, achieving 100% sensitivity and 97.3% specificity with 66 clinical samples. Our instrument offers a cost-effective (USD 100) solution with automated result interpretation and superior sensitivity compared to visual inspection. While the prototype was tested with SARS-CoV-2 RNA samples, its versatility extends to detecting other pathogens using alternative primers, showcasing its potential for broader applications in biosensing.

A low-cost PCR instrument for molecular disease diagnostics based on customized printed circuit board heaters.

This article describes the fabrication of a low-cost Polymerase Chain Reaction (PCR) instrument to detect diseases. In order to reduce the instrument price and simplify construction we developed an alternative fabrication process, transforming conventional printed circuit boards (PCB) in heating elements, avoiding the use of aluminum heating/cooling blocks and Peltier devices. To cool down the reaction a simple computer fan was used. The vial holder was fabricated using two double side PCB boards assembled in a sandwich-like configuration. The bottom PCB has a resistance of 0.9 Ω used to heat the reaction mix, while the top layer has a resistance of 1.1 Ω to heat the vial body, preventing vapor condensation. The top board was maintained at ~ 110 ± 1 °C during all cycles. The final device was able to heat and cool down the reaction at rates of ~ 2.0 °C/s, a rate comparable to commercial thermocyclers. An SMD NTC thermistor was used as temperature sensors, and a PID (proportional-integral-derivative) control algorithm was implemented to acquire and precisely control the temperature. We also discuss how the instrument is calibrated. The device was tested successfully for the amplification of T. pallidum (Syphilis) bacterial DNA and Zika virus RNA samples, showing similar performance to a commercial PCR instrument.