ABSTRACT
The development of sensitive and affordable testing devices for infectious diseases is essential to preserve public health, especially in pandemic scenarios. In this work, we have developed an attractive analytical method to monitor products of genetic amplification, particularly the loop-mediated isothermal amplification reaction (RT-LAMP). The method is based on electrochemical impedance measurements and the distribution of relaxation times model, to provide the so-called time-constant-domain spectroscopy (TCDS). The proposed method is tested for the SARS-CoV-2 genome, since it has been of worldwide interest due to the COVID-19 pandemic. Particularly, once the method is calibrated, its performance is demonstrated using real wastewater samples. Moreover, we propose a simple classification algorithm based on TCDS data to discriminate among positive and negative samples. Results show how a TCDS-based method provides an alternative mechanism for label-free and automated assays, exhibiting robustness and specificity for genetic detection.
ABSTRACT
The current pandemic COVID-19 caused by the coronavirus SARS-CoV-2, has generated different economic, social and public health problems. Moreover, wastewater-based epidemiology could be a predictor of the virus rate of spread to alert on new outbreaks. To assist in epidemiological surveillance, this work introduces a simple, low-cost and affordable electrochemical sensor to specifically detect N and ORF1ab genes of the SARS-CoV-2 genome. The proposed sensor works based on screen-printed electrodes acting as a disposable test strip, where the reverse transcription loop-mediated isothermal amplification (RT-LAMP) reaction takes place. Electrochemical detection relies upon methylene blue as a redox intercalator probe, to provide a diffusion-controlled current encoding the presence and concentration of RT-LAMP products, namely amplicons or double-stranded DNA. We test the performance of the sensor by testing real wastewater samples using end-point and time course measurements. Results show the ability of the electrochemical test strip to specifically detect and quantify RT-LAMP amplicons below to ~ 2.5 × 10-6 ng/µL exhibiting high reproducibility. In this sense, our RT-LAMP electrochemical sensor is an attractive, efficient and powerful tool for rapid and reliable wastewater-based epidemiology studies.
ABSTRACT
The COVID-19 pandemic caused by the coronavirus SARS-CoV-2 has proven the need for developing reliable and affordable technologies to detect pathogens. Particularly, the detecting the genome in wastewater could be an indicator of the transmission rate to alert on new outbreaks. However, wastewater-based epidemiology remains a technological challenge to develop affordable technologies for sensing pathogens. In this work, we introduce a label-free and portable field-effect transistor (FET)-based sensor to detect N and ORF1ab genes of the SARS-CoV-2 genome. Our sensor integrates the reverse transcription loop-mediated isothermal amplification (RT-LAMP) reaction as a cost-effective molecular detection exhibiting high specificity. The detection relies upon pH changes, due to the RT-LAMP reaction products, which are detected through a simple, but effective, extended-gate FET sensor (EGFET). We evaluate the proposed device by measuring real wastewater samples to detect the presence of SARS-CoV-2 genome, achieving a limit of detection of 0.31 × 10−3 ng/ μ L for end-point measurement. Moreover, we find the ability of the sensor to perform real-time-like analysis, showing that the RT-LAMP reaction provides a good response after 15 min for concentrations as low as 0.37 ng/ μ L. Hence, we show that our EGFET sensor offers a powerful tool to detect the presence of the SARS-CoV-2 genome with a naked-eye method, in a straightforward way than the conventional molecular methods for wastewater analysis. [Display omitted] • Label-free extended-gate field-effect transistor sensor for detect SARS-CoV-2 genome. • Portable and reliable sensing based on isothermal amplification reaction. • Detection and quantification of nucleic-acids in real wastewater samples. • End-pint and time course detection of RT-LAMP products. • The wastewater-based epidemiology can use this method in limited-resource conditions. [ FROM AUTHOR]
ABSTRACT
The COVID-19 pandemic caused by the coronavirus SARS-CoV-2 has proven the need for developing reliable and affordable technologies to detect pathogens. Particularly, the detecting the genome in wastewater could be an indicator of the transmission rate to alert on new outbreaks. However, wastewater-based epidemiology remains a technological challenge to develop affordable technologies for sensing pathogens. In this work, we introduce a label-free and portable field-effect transistor (FET)-based sensor to detect N and ORF1ab genes of the SARS-CoV-2 genome. Our sensor integrates the reverse transcription loop-mediated isothermal amplification (RT-LAMP) reaction as a cost-effective molecular detection exhibiting high specificity. The detection relies upon pH changes, due to the RT-LAMP reaction products, which are detected through a simple, but effective, extended-gate FET sensor (EGFET). We evaluate the proposed device by measuring real wastewater samples to detect the presence of SARS-CoV-2 genome, achieving a limit of detection of 0.31 × 10−3 ng/μL for end-point measurement. Moreover, we find the ability of the sensor to perform real-time-like analysis, showing that the RT-LAMP reaction pro-vides a good response after 15 min for concentrations as low as 0.37 ng/μL. Hence, we show that our EGFET sensor offers a powerful tool to detect the presence of the SARS- CoV-2 genome with a naked-eye method, in a straightforward way than the conventional molecular methods for wastewater analysis.