scholarly journals Portable and Label-Free Quantitative Loop-Mediated Isothermal Amplification (LF-qLamp) for Reliable COVID-19 Diagnostics in Three Minutes of Reaction Time: Arduino-Based Detection System Assisted by a pH Microelectrode

Biosensors ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 386
Author(s):  
Mario Moisés Alvarez ◽  
Sergio Bravo-González ◽  
Everardo González-González ◽  
Grissel Trujillo-de Santiago
Keyword(s):  
Real Time ◽  
Detection System ◽  
Genetic Material ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Label Free ◽  
Viral Pathogens ◽  
Loop Mediated Isothermal Amplification ◽  
Amplification System

Loop-mediated isothermal amplification (LAMP) has been recently studied as an alternative method for cost-effective diagnostics in the context of the current COVID-19 pandemic. Recent reports document that LAMP-based diagnostic methods have a comparable sensitivity and specificity to that of RT-qPCR. We report the use of a portable Arduino-based LAMP-based amplification system assisted by pH microelectrodes for the accurate and reliable diagnosis of SARS-CoV-2 during the first 3 min of the amplification reaction. We show that this simple system enables a straightforward discrimination between samples containing or not containing artificial SARS-CoV-2 genetic material in the range of 10 to 10,000 copies per 50 µL of reaction mix. We also spiked saliva samples with SARS-CoV-2 synthetic material and corroborated that the LAMP reaction can be successfully monitored in real time using microelectrodes in saliva samples as well. These results may have profound implications for the design of real-time and portable quantitative systems for the reliable detection of viral pathogens including SARS-CoV-2.

scholarly journals Portable and label-free quantitative Loop-mediated Isothermal Amplification (qLAMP) for reliable COVID-19 diagnostics in 3 minutes: An Arduino-based detection system assisted by a pH microelectrode

2021 ◽  
Author(s):  
Mario Moisés Alvarez ◽  
Sergio Bravo-González ◽  
Everardo González-González ◽  
Grissel Trujillo-de Santiago
Keyword(s):  
Real Time ◽  
Detection System ◽  
Genetic Material ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Label Free ◽  
Viral Pathogens ◽  
Loop Mediated Isothermal Amplification ◽  
Amplification System

Loop-mediated isothermal amplification (LAMP) has been recently studied as an alternative method for cost-effective diagnostics in the context of the current COVID-19 pandemic. Recent reports document that LAMP-based diagnostic methods have a comparable sensitivity and specificity to that of RT-qPCR. We report the use of a portable Arduino-based LAMP-based amplification system assisted by pH microelectrodes for the accurate and reliable diagnosis of SARS-CoV-2 during the first 3 minutes of the amplification reaction. We show that this simple system enables a straightforward discrimination between samples containing or not containing artificial SARS-CoV-2 genetic material in the range of 10 to 10,000 copies per 50 μL of reaction mix. We also spiked saliva samples with SARS-CoV-2 synthetic material and corroborated that the LAMP reaction can be successfully monitored in real time using microelectrodes in saliva samples as well. These results may have profound implications for the design of real-time and portable quantitative systems for the reliable detection of viral pathogens including SARS-CoV-2.

scholarly journals Comparison of Loop-Mediated Isothermal Amplification and Real-Time PCR Assays for Detection ofStrongyloidesLarvae in Different Specimen Matrices

2019 ◽  
Vol 57 (4) ◽  
Author(s):  
Matthew R. Watts ◽  
Rady Kim ◽  
Vishal Ahuja ◽  
Gemma J. Robertson ◽  
Yasmin Sultana ◽  
...  
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Chronic Infection ◽  
Lamp Assay ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Percentage Agreement ◽  
Content Type ◽  
Loop Mediated Isothermal Amplification ◽  
Stool Specimens

ABSTRACTStrongyloides stercoraliscan cause disease that ranges from asymptomatic chronic infection to fatal hyperinfection. Diagnosis from stool can be challenging because the most sensitive conventional tests require live larvae to be effective and there can be low larval output in chronic infection. Nucleic acid amplification tests (NAAT) have been developed to complement existing diagnostic methods. We compared a recently developed loop-mediated isothermal amplification (LAMP) assay with a real-time PCR that has previously been validated with larval microscopy. The limits of detection—quantified using serial dilutions of DNA extracts from singleStrongyloides rattithird-stage (L3) larvae spiked into approximately 250 µl of 5 differentS. stercoralis-negative stool specimens—were 10−3(1/5 replicates) and 10−2(1/5 replicates) dilutions for PCR and LAMP, respectively. PCR was positive for 4/5 replicates at 10−2. LAMP was compared to PCR using extracts from 396 stool specimens collected in Bangladesh and Australia, of which 53 were positive and 343 were negative by PCR. The positive percentage agreement of LAMP was 77.3% (95% score confidence interval [CI], 64.5 to 86.6). The negative percentage agreement was 100% (95% CI, 98.9 to 100). In a preliminary investigation, PCR and LAMP assays were positive using DNA extracted from serum (PCR, 3/16 extracts; LAMP, 2/16 extracts) and bronchoalveolar lavage fluid (PCR and LAMP, 2/2 extracts), demonstrating proof of concept. Compared to PCR, the lower number of positive results using the LAMP assay may have been due to reaction inhibitors and DNA degradation, and strategies to improve the LAMP assay are discussed.

scholarly journals Rapid and Sensitive Detection of Tomato Brown Rugose Fruit Virus in Tomato and Pepper Seeds by Reverse Transcription Loop-Mediated Isothermal Amplification Assays (Real Time and Visual) and Comparison With RT-PCR End-Point and RT-qPCR Methods

2021 ◽  
Vol 12 ◽  
Author(s):  
Domenico Rizzo ◽  
Daniele Da Lio ◽  
Alessandra Panattoni ◽  
Chiara Salemi ◽  
Giovanni Cappellini ◽  
...  
Keyword(s):  
Real Time ◽  
Reverse Transcription ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
The European Union ◽  
Rt Pcr ◽  
Loop Mediated Isothermal Amplification ◽  
End Point ◽  
Specificity And Sensitivity ◽  
The Status

Tomato brown rugose fruit virus (ToBRFV) represents an emerging viral threat to the productivity of tomato and pepper protected cultivation worldwide. This virus has got the status of quarantine organism in the European Union (EU) countries. In particular, tomato and pepper seeds will need to be free of ToBRFV before entering the EU and before coming on the market. Thus, lab tests are needed. Here, we develop and validate a one-step reverse transcription LAMP platform for the detection of ToBRFV in tomato and pepper leaves, by real-time assay [reverse transcription loop-mediated isothermal amplification (RT-LAMP)] and visual screening (visual RT-LAMP). Moreover, these methods can also be applied successfully for ToBRFV detection in tomato and pepper seeds. The diagnostic specificity and sensitivity of both RT-LAMP and visual RT-LAMP are both 100%, with a detection limit of nearly 2.25 fg/μl, showing the same sensitivity as RT-qPCR Sybr Green, but 100 times more sensitive than end-point RT-PCR diagnostic methods. In artificially contaminated seeds, the proposed LAMP assays detected ToBRFV in 100% of contaminated seed lots, for up to 0.025–0.033% contamination rates in tomato and pepper, respectively. Our results demonstrate that the proposed LAMP assays are simple, inexpensive, and sensitive enough for the detection of ToBRFV, especially in seed health testing. Hence, these methods have great potential application in the routine detection of ToBRFV, both in seeds and plants, reducing the risk of epidemics.

scholarly journals Loop-Mediated Isothermal Amplification (LAMP): A Rapid, Sensitive, Specific, and Cost-Effective Point-of-Care Test for Coronaviruses in the Context of COVID-19 Pandemic

Biology ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 182 ◽  
Author(s):  
Robin Augustine ◽  
Anwarul Hasan ◽  
Suvarthi Das ◽  
Rashid Ahmed ◽  
Yasuyoshi Mori ◽  
...  
Keyword(s):  
Low Income ◽  
Point Of Care ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
Diagnostic Methods ◽  
Diagnostic Tools ◽  
Paper Strip ◽  
Ongoing Research ◽  
Loop Mediated Isothermal Amplification ◽  
Point Of Care Test

The rampant spread of COVID-19 and the worldwide prevalence of infected cases demand a rapid, simple, and cost-effective Point of Care Test (PoCT) for the accurate diagnosis of this pandemic. The most common molecular tests approved by regulatory bodies across the world for COVID-19 diagnosis are based on Polymerase Chain Reaction (PCR). While PCR-based tests are highly sensitive, specific, and remarkably reliable, they have many limitations ranging from the requirement of sophisticated laboratories, need of skilled personnel, use of complex protocol, long wait times for results, and an overall high cost per test. These limitations have inspired researchers to search for alternative diagnostic methods that are fast, economical, and executable in low-resource laboratory settings. The discovery of Loop-mediated isothermal Amplification (LAMP) has provided a reliable substitute platform for the accurate detection of low copy number nucleic acids in the diagnosis of several viral diseases, including epidemics like Severe Acute Respiratory Syndrome (SARS) and Middle East Respiratory Syndrome (MERS). At present, a cocktail of LAMP assay reagents along with reverse transcriptase enzyme (Reverse Transcription LAMP, RT-LAMP) can be a robust solution for the rapid and cost-effective diagnosis for COVID-19, particularly in developing, and low-income countries. In summary, the development of RT-LAMP based diagnostic tools in a paper/strip format or the integration of this method into a microfluidic platform such as a Lab-on-a-chip may revolutionize the concept of PoCT for COVID-19 diagnosis. This review discusses the principle, technology and past research underpinning the success for using this method for diagnosing MERS and SARS, in addition to ongoing research, and the prominent prospect of RT-LAMP in the context of COVID-19 diagnosis.

Low-Cost Label-Free Bio-Detection System Using Double-Sided Grating Waveguide Couplers

2015 ◽  
Author(s):  
Yi-Fan Ku ◽  
Hsun-Yuan Li ◽  
Yu-Chung Lin ◽  
Wen-Hsin Hsieh ◽  
Guo-En Chang
Keyword(s):  
Real Time ◽  
Detection System ◽  
Light Emitting Diode ◽  
Low Cost ◽  
Cost Effective ◽  
High Refractive Index ◽  
Optical Intensity ◽  
Label Free ◽  
Light Emitting ◽  
Label Free Detection

Low-cost label-free bio-sensing systems have long been desired to enable rapid, sensitive, quantitative, and high-throughput biosensing for bio-medical and chemical applications. Here we present an optical bio-detection system consists of injection-molded biosensors based on double-sided grating waveguide couplers and an optical intensity-based detection platform for low-cost, real-time, and label-free biosensing. The biosensors were fabricated combining injection-molding and sputtering techniques, providing unique advantages of low-cost and reduced production time. A simple and cost-effective optical intensity-based detection system employing a low-cost light emitting diode and a simple photodetector is also developed to perform label-free biosensing. We demonstrate that a high refractive index resolution of 6.43 × 10−5 RIU is achieved with this compact bio-sensing system, showing great promises for low-cost, real-time, label-free detection in bio-medical and chemical applications.

scholarly journals Development of a Novel Loop Mediated Isothermal Amplification Assay (LAMP) for the Rapid Detection of Epizootic Haemorrhagic Disease Virus

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2187
Author(s):  
Paulina Rajko-Nenow ◽  
Emma L. A. Howson ◽  
Duncan Clark ◽  
Natasha Hilton ◽  
Aruna Ambagala ◽  
...  
Keyword(s):  
Real Time ◽  
Rapid Detection ◽  
High Throughput Screening ◽  
Limit Of Detection ◽  
Lamp Assay ◽  
Cost Effective ◽  
Virus Genome ◽  
Isothermal Amplification ◽  
Loop Mediated Isothermal Amplification ◽  
Haemorrhagic Disease

Epizootic haemorragic disease (EHD) is an important disease of white-tailed deer and can cause a bluetongue-like illness in cattle. A definitive diagnosis of EHD relies on molecular assays such as real-time RT-qPCR or conventional PCR. Reverse transcription loop-mediated isothermal amplification (RT-LAMP) is a cost-effective, specific, and sensitive technique that provides an alternative to RT-qPCR. We designed two sets of specific primers targeting segment-9 of the EHD virus genome to enable the detection of western and eastern topotypes, and evaluated their performance in singleplex and multiplex formats using cell culture isolates (n = 43), field specimens (n = 20), and a proficiency panel (n = 10). The limit of detection of the eastern and western RT-LAMP assays was estimated as ~24.36 CT and as ~29.37 CT in relation to real-time RT-qPCR, respectively, indicating a greater sensitivity of the western topotype singleplex RT-LAMP. The sensitivity of the western topotype RT-LAMP assay, relative to the RT-qPCR assay, was 72.2%, indicating that it could be theoretically used to detect viraemic cervines and bovines. For the first time, an RT-LAMP assay was developed for the rapid detection of the EHD virus that could be used as either a field test or high throughput screening tool in established laboratories to control the spread of EHD.

scholarly journals The detection of SARS-CoV-2 using reverse transcription loop-mediated isothermal amplification (RT-LAMP) in developing country

10.51511/pr.9 ◽  
2021 ◽  
Vol 1 (1) ◽  
pp. 9
Author(s):  
Muhammad Rizky Wibowo ◽  
Erna Harfiani ◽  
Sarmoko Sarmoko ◽  
Yudhi Nugraha
Keyword(s):  
Reverse Transcription ◽  
Rural Areas ◽  
Genetic Material ◽  
Cost Effective ◽  
Isothermal Amplification ◽  
World Health ◽  
Rt Pcr ◽  
Loop Mediated Isothermal Amplification ◽  
Alternative Detection ◽  
Health Organization

The severe acute respiratory syndrome coronavirus (SARS-CoV-2) has infected the human system resulting in Covid-19, and has spread rapidly worldwide. Therefore, a fast, simple, cost-effective, and accurate detecting tool is required. The standard diagnostic tool of the World Health Organization is the reverse transcription-polymerase chain reaction (RT-PCR). This method detects the presence of viral genetic material in the human body with accurate results. However, it has several limitations in terms of equipment, personnel, duration, and cost. Therefore, a fast, simple, and sensitive alternative detection, is required, one of which is the reverse transcription loop-mediated isothermal amplification (RT-LAMP) that functions under isothermal conditions. This method is battery-driven, hence, easy to move closer to the patient. Conclusively, the RT-LAMP test for SARS CoV-2 diagnosis produces comparable sensitivity to a standard RT-PCR and is more suitable for resource-poor settings, such as rural areas of developing countries.

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