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

Huynh Quoc Nguyen,Tae Seok Seo,Hau Van Nguyen,Van Dan Nguyen

doi:10.1038/s41598-020-72095-3

Huynh Quoc Nguyen, Tae Seok Seo + Show 2 more

Open Access

https://doi.org/10.1038/s41598-020-72095-3

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Journal: Scientific Reports	Publication Date: Sep 15, 2020
Citations: 50	License type: open-access

Affiliation: Kyung Hee University

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.

Highlights

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
For the quantitative as well as qualitative analysis of the pathogens using the colorimetric loop-mediated isothermal amplification (LAMP) reaction, we developed an integrated gene box (i-Genbox), which consists of a commercial smartphone, an auxiliary USB battery (5 V), a LAMP reaction box (6 cm height × 5 cm width × 8 cm length) equipped with a white LED, a LAMP chip, a DC converter, a film heater (Fig. 1A,B)
At a voltage of 24.7 V, the temperature of the film heater was maintained at 65 °C that is adequate for LAMP reaction

Summary

Introduction

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. The colorimetric detection is to use a metal indicator dye, for example, Eriochrome Black T (EBT) and hydroxy naphthol blue (HNB), or a pH-sensitive dye such as phenol red and NeuRed in the reaction cocktail, and the target gene amplification can be monitored by observing the color of the reaction mixture without need of laboratory-based instruments or expensive detectors/sensors. Considering all these factors, we believe that the best combination for the POCT is the isothermal amplification with the colorimetric detection. We addressed the above requirements to propose a smartphone integrated POCT platform for nucleic acid quantification with a new colorimetric approach, simplicity of production and operation and a mobile app

Methods

Results

Conclusion