Quantification of colorimetric isothermal amplification on the smartphone and its open-source app for point-of-care pathogen detection

Abstract

The increasing risk of infectious pathogens, especially in the under-developed countries, is demanding the development of point-of-care (POC) nucleic acid testing in the low-resource setting conditions. Here, we describe a methodology for colorimetric quantitative analysis of nucleic acid using an easy-to-build smartphone-based platform, offering low-cost, portability, simplicity in operation, and user-friendliness. The whole system consists of a hand-held box equipped with a smartphone, a film heater, a white LED, a loop-mediated isothermal amplification (LAMP) chip, and a DC converter, and all the processes were powered by a portable battery of 5 V. Upon the amplification of the target gene by an Eriochrome Black T-mediated LAMP reaction, the color of the LAMP reaction was changed from violet to blue that was real-time recorded by a smartphone camera. To keep track of the progress of the color change, we developed a novel mobile app in which a hue value was accepted as an indicator for color transition and for determining the threshold time of the amplification reaction. A calibration curve could be generated by plotting the logarithm of the known concentration of the DNA templates versus the threshold time, and it can be used to predict the copy number of nucleic acids in the test samples. Thus, the proposed mobile platform can inform us of not only qualitative but also quantitative results of the pathogens. We believe that this advanced colorimetric approach and the mobile app can expand the potentials of the smartphone for the future POCT system in the bio-diagnostic fields.

Loop-mediated isothermal amplification (LAMP) assay targeting RLEP for detection of Mycobacterium leprae in leprosy patients

“…The addition of UDG can eliminate cross-contamination, the potential for which is further limited by having experienced personnel operate the assay in a point-of-care setting. Recent studies have successfully utilized pHbased dyes for better visualization of results (Yuan, Kong et al 2018;Rabe and Cepko, 2020), and smartphone-based quantification of colorimetric results has also been shown to aid interpretation of results (Nguyen et al, 2020;Yin, Pandian et al 2020). In this study, all conditions were optimized to improve the sensitivity and specificity of detection.…”

Toward a next-generation diagnostic tool: A review on emerging isothermal nucleic acid amplification techniques for the detection of SARS-CoV-2 and other infectious viruses

“…Real-time and quantitative LAMP with a colorimetric readout mode are also possible. One such study used smartphones and open-source apps to monitor the dye Eriochrome Black T color change in real time in a miniaturized LAMP reaction [ 80 ]. A similar real-time colorimetric LAMP reaction was performed in a miniaturized platform using smartphone-based color detection of the phenol red indicator [ 81 ].…”

Recent advances and challenges of biosensing in point-of-care molecular diagnosis

“…The smartphone can realize wireless connectivity with the biosensing system through blue tooth or a WIFI interface for device control, data collection, display, and analysis [182 ] . It can also serve to capture the image of the reaction products and communicate with cloud-connected computing systems for further image processing by professional software or algorithms [183] , [184] , [185] . In particular, the colorimetric assay-based biosensing systems are often combined with a smartphone to capture the result image and analyze the hue value of RGB channels, which can provide a more promising quantitative conclusion compared with naked eyes.…”

“…In particular, the colorimetric assay-based biosensing systems are often combined with a smartphone to capture the result image and analyze the hue value of RGB channels, which can provide a more promising quantitative conclusion compared with naked eyes. For instance, Nguyen et al integrated a smartphone into a pathogen detection system and designed a mobile app for real-time recording and processing using the hue value as the indicator of color transition [184 ] . Furthermore, as machine learning and deep learning technologies flourish, it's a convincing and reliable way to combine biosensing system with artificial intelligence by a smartphone to improve sensitivity and specificity of molecular diagnosis testing [186 ] .…”