ArticleNucleic acids research2016
Lack of correlation between reaction speed and analytical sensitivity in isothermal amplification reveals the value of digital methods for optimization: validation using digital real-time RT-LAMP.
Article in Nucleic acids research, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 15 papers.
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Who cites it
15 citing papers in PubMed.
- Article
- Digital droplet RT-LAMP increases speed of SARS-CoV-2 viral RNA detection.Smart medicine · 2024Article
- Quantitative analysis of miRNAs using SplintR ligase-mediated ligation of complementary-pairing probes enhanced by RNase H (SPLICER)-qPCR.Molecular therapy. Nucleic acids · 2023Article
- Review
- Polymerase chain reaction and loop-mediated isothermal amplification targeting lic13162, lic20239, and lipL32 genes for leptospirosis diagnosis.Brazilian journal of microbiology : [publication of the Brazilian Society for Microbiology] · 2022Article
- Loop-Mediated Isothermal Amplification: From Theory to Practice.Russian journal of bioorganic chemistry · 2022Article
- Rapid, Affordable, and Scalable SARS-CoV-2 Detection From Saliva.Journal of biomolecular techniques : JBT · 2021Article
- Loop-Mediated Isothermal Amplification Detection of SARS-CoV-2 and Myriad Other Applications.Journal of biomolecular techniques : JBT · 2021Review
- ADiagnostics (Basel, Switzerland) · 2021Article
- Development of Reverse Transcription Loop-Mediated Isothermal Amplification Assays Targeting Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2).The Journal of molecular diagnostics : JMD · 2020Article
- Real-time kinetics and high-resolution melt curves in single-molecule digital LAMP to differentiate and study specific and non-specific amplification.Nucleic acids research · 2020Article
- Quantitative and rapid Plasmodium falciparum malaria diagnosis and artemisinin-resistance detection using a CMOS Lab-on-Chip platform.Biosensors & bioelectronics · 2019Article
- Slip-driven microfluidic devices for nucleic acid analysis.Biomicrofluidics · 2019Review
- Real-Time, Digital LAMP with Commercial Microfluidic Chips Reveals the Interplay of Efficiency, Speed, and Background Amplification as a Function of Reaction Temperature and Time.Analytical chemistry · 2019Article
- Instrument for Real-Time Digital Nucleic Acid Amplification on Custom Microfluidic Devices.PloS one · 2016Article
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Authors and funding
4 authors.
Funding
Abstract
In this paper, we asked if it is possible to identify the best primers and reaction conditions based on improvements in reaction speed when optimizing isothermal reactions. We used digital single-molecule, real-time analyses of both speed and efficiency of isothermal amplification reactions, which revealed that improvements in the speed of isothermal amplification reactions did not always correlate with improvements in digital efficiency (the fraction of molecules that amplify) or with analytical sensitivity. However, we observed that the speeds of amplification for single-molecule (in a digital device) and multi-molecule (e.g. in a PCR well plate) formats always correlated for the same conditions. Also, digital efficiency correlated with the analytical sensitivity of the same reaction performed in a multi-molecule format. Our finding was supported experimentally with examples of primer design, the use or exclusion of loop primers in different combinations, and the use of different enzyme mixtures in one-step reverse-transcription loop-mediated amplification (RT-LAMP). Our results show that measuring the digital efficiency of amplification of single-template molecules allows quick, reliable comparisons of the analytical sensitivity of reactions under any two tested conditions, independent of the speeds of the isothermal amplification reactions.
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