Browse
Submit Data
Databases
API
Help

Dataset Information

16 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Towards a versatile and economic Chagas Disease point-of-care testing system, by integrating loop-mediated isothermal amplification and contactless/label-free conductivity detection.

ABSTRACT: Rapid diagnosis by using small, simple, and portable devices could represent one of the best strategies to limit the damage and contain the spread of viral, bacterial or protozoa diseases, principally when they can be transmitted by air and are highly contagious, as some respiratory viruses are. The presence of antibodies in blood or serum samples is not the best option for deciding when a person must be quarantined to stop transmission of disease, given that cured patients have antibodies, so the best diagnosis methods rely on the use of nucleic acid amplification procedures. Here we present a very simple device and detection principle, based on paper discs coupled to contactless conductivity (C4D) sensors, can provide fast and easy diagnostics that are needed when an epidemic outbreak develops. The paper device presented here solves one of the main drawbacks that nucleic acid amplification tests have when they are performed outside of central laboratories. As the device is sealed before amplification and integrally disposed in this way, amplimers release cannot occur, allowing repetitive testing in the physician's practice, ambulances, or other places that are not prepared to avoid cross-contamination of new samples. The use of very low volume samples allows efficient reagent use and the development of low cost, simple, and disposable point-of-care diagnostic systems.

SUBMITTER: Figueredo F

PROVIDER: S-EPMC8153438 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Developments in integrating nucleic acid isothermal amplification and detection systems for point-of-care diagnostics.

Project description:Early disease detection through point-of-care (POC) testing is vital for quickly treating patients and preventing the spread of harmful pathogens. Disease diagnosis is generally accomplished using quantitative polymerase chain reaction (qPCR) to amplify nucleic acids in patient samples, permitting detection even at low target concentrations. However, qPCR requires expensive equipment, trained personnel, and significant time. These resources are not available in POC settings, driving researchers to instead utilize isothermal amplification, conducted at a single temperature, as an alternative. Common isothermal amplification methods include loop-mediated isothermal amplification, recombinase polymerase amplification, rolling circle amplification, nucleic acid sequence-based amplification, and helicase-dependent amplification. There has been a growing interest in combining such amplification methods with POC detection methods to enable the development of diagnostic tests that are well suited for resource-limited settings as well as developed countries performing mass screenings. Exciting developments have been made in the integration of these two research areas due to the significant impact that such approaches can have on healthcare. This review will primarily focus on advances made by North American research groups between 2015 and June 2020, and will emphasize integrated approaches that reduce user steps, reliance on expensive equipment, and the system's time-to-result.

| S-EPMC7529604 | biostudies-literature

Hinge-initiated Primer-dependent Amplification of Nucleic Acids (HIP) - A New Versatile Isothermal Amplification Method.

Project description:The growing demand for cost-effective nucleic acid detection assays leads to an increasing number of different isothermal amplification reaction methods. However, all of the most efficient methods suffer from highly complex assay conditions due to the use of complicated primer sets and/or auxiliary enzymes. The present study describes the application of a new linker moiety that can be incorporated between a primer and a secondary target binding site which can act both as a block to polymerase extension as well as a hinge for refolding. This novel "hinge-primer" approach results in an efficient regeneration of the primer binding site and thus improves the strand-displacement and amplification process under isothermal conditions. Our investigations revealed that the reaction with forward and reverse hinge-primer including an abasic site is very efficient. The assay complexity can be reduced by combining the hinge-primer with a corresponding linear primer. Furthermore, the reaction speed can be increased by reducing the length of the amplified target sequence. We tested the sensitivity down to 104 copies and found a linear correlation between reaction time and input copy number. Our approach overcomes the usually cumbersome primer-design and extends the range of isothermal amplification methods using a polymerase with strand-displacement activity.

| S-EPMC5550438 | biostudies-literature

Amplicon Competition Enables End-Point Quantitation of Nucleic Acids Following Isothermal Amplification.

Project description:It is inherently difficult to quantitate nucleic acid analytes with most isothermal amplification assays. We developed loop-mediated isothermal amplification (LAMP) reactions in which competition between defined numbers of "false" and "true" amplicons leads to order of magnitude quantitation by a single endpoint determination. These thresholded LAMP reactions were successfully used to directly and quantitatively estimate the numbers of nucleic acids in complex biospecimens, including directly from cells and in sewage, with the values obtained closely correlating with qPCR quantitations. Thresholded LAMP reactions are amenable to endpoint readout by cell phone, unlike other methods that require continuous monitoring, and should therefore prove extremely useful in developing one-pot reactions for point-of-care diagnostics without needing sophisticated material or informatics infrastructure.

| S-EPMC5890436 | biostudies-literature

Label-free detection of microRNA based on coupling multiple isothermal amplification techniques.

Project description:MicroRNA (miRNA) was a promising class of cancer biomarkers. Here we developed a label-free method for sensitive measurement of let-7d miRNA based on multiple amplification techniques. The primer will bind to the duplex strand DNA that was formed by stem-loop template and target let-7d to initiate strand displacement amplification (SDA) in tandem. The released single strand DNA will be a primer to bind the circular template to initiate rolling circle amplification (RCA). The products based on multiple amplifications will be detected by a standard fluorimeter with N-methyl mesoporphyrin IX (NMM) as the fluorescent indicator. The proposed method exhibited excellent selectivity and high sensitivity with a detection limit of as low as 1.5?×?10-13?M. Moreover, this methodology was used for the determination of biomolecules in real serum samples with satisfying results.

| S-EPMC5078768 | biostudies-literature

A versatile isothermal amplification assay for the detection of leptospires from various sample types.

Project description:BackgroundLeptospirosis is a zoonotic disease caused by bacteria of the genus Leptospira that affects both humans and animals worldwide. Early detection of the pathogen in humans is crucial for early intervention and control of the progression of the disease to a severe state. It is also vitally important to be able to detect the presence of the pathogen in carrier animals to control the spread of the disease from the environment. Here we developed a simple and rapid loop-mediated isothermal amplification (LAMP) assay targeting the leptospiral secY gene.ResultsSeveral reaction conditions of the LAMP reaction were optimized to ensure efficient amplification of the target DNA. The sensitivity of the developed LAMP assay obtained using a pure Leptospira culture was 2 × 104 copies of genomic DNA per reaction (equivalent to 0.1 ng) for a 40-minute reaction time. No cross-reactions were observed in the LAMP reaction against a series of non-leptospiral bacteria, indicating a specific reaction. The applicability of the LAMP assay was demonstrated on human blood and urine specimens collected from suspected leptospirosis patients and rat kidney specimens collected from suspected leptospirosis outbreak areas and high-risk areas. The developed LAMP assay demonstrated a higher detection rate for leptospiral DNA compared with the polymerase chain reaction (PCR) assay, possibly due to the presence of inhibitory substances, especially in rat kidney specimens, to which the PCR method is more susceptible. The present findings also highlight the importance of urine sample collection from patients for routine monitoring of the disease.ConclusionsIn short, the developed LAMP assay can serve as a feasible alternative tool for the diagnosis of leptospirosis and be used for epidemiological and environmental surveillance of the disease, considering its robustness, rapidity, sensitivity, and specificity, as demonstrated in this study.

| S-EPMC8918162 | biostudies-literature

Label-Free Isothermal Amplification Assay for Specific and Highly Sensitive Colorimetric miRNA Detection.

Project description:We describe a new method for the detection of miRNA in biological samples. This technology is based on the isothermal nicking enzyme amplification reaction and subsequent hybridization of the amplification product with gold nanoparticles and magnetic microparticles (barcode system) to achieve naked-eye colorimetric detection. This platform was used to detect a specific miRNA (miRNA-10b) associated with breast cancer, and attomolar sensitivity was demonstrated. The assay was validated in cell culture lysates from breast cancer cells and in serum from a mouse model of breast cancer.

| S-EPMC5046170 | biostudies-literature

Bacterial Concentration Detection using a PCB-based Contactless Conductivity Sensor.

Project description:Capacitively coupled contactless conductivity detection (C?D) is an improved approach to avoid the problems of labor-intensive, time-consuming and insufficient accuracy of plate count as well as the high-cost apparatus of flow cytometry (FCM) in bacterial counting. This article describes a novel electrode-integrated printed-circuit-board (PCB)-based C?D device, which supports the simple and safe exchange of capillaries and improves the sensitivity and repeatability of the contactless detection. Furthermore, no syringe pump is needed in the detection, it reduces the system size, and, more importantly, avoids the effect on the bacteria due to high pressure. The recovered bacteria after C?D detection at excitation of 25 Vpp and 60?120 kHz were analyzed by flow cytometry, and a survival rate higher than 96% was given. It was verified that C?D detection did not influence the bacterial viability. Moreover, bacteria concentrations from 10? cells/mL to 10? cells/mL were measured in a linear range, and relative standard deviation (RSD) is below 0.2%. In addition, the effects on bacteria and C?D from background solutions were discussed. In contrast to common methods used in most laboratories, this method may provide a simple solution to in situ detection of bacterial cultures.

| S-EPMC6356519 | biostudies-literature

Sensitive electrochemical biosensor combined with isothermal amplification for point-of-care COVID-19 tests.

Project description:We report an electrochemical biosensor combined with recombinase polymerase amplification (RPA) for rapid and sensitive detection of severe acute respiratory syndrome coronavirus 2. The electrochemical biosensor based on a multi-microelectrode array allows the detection of multiple target genes by differential pulse voltammetry. The RPA reaction involves hybridization of the RPA amplicon with thiol-modified primers immobilized on the working electrodes, which leads to a reduction of current density as amplicons accumulate. The assay results in shorter "sample-to-answer" times than conventional PCR without expensive thermo-cycling equipment. The limits of detection are about 0.972 fg/μL (RdRP gene) and 3.925 fg/μL (N gene), which are slightly lower than or comparable to that of RPA assay results obtained by gel electrophoresis without post-amplification purification. The combination of electrochemical biosensors and the RPA assay is a rapid, sensitive, and convenient platform that can be potentially used as a point-of-care test for the diagnosis of COVID-19.

| S-EPMC7970423 | biostudies-literature

Evaluation and improvement of isothermal amplification methods for point-of-need plant disease diagnostics.

Project description:A number of isothermal DNA amplification technologies claim to be ideal for point-of-need (PON) applications as they enable reactions to be performed using a single-temperature heat source (e.g. water bath). Thus, we examined several isothermal amplification methods focusing on simplicity, cost, sensitivity and reproducibility to identify the most suitable method(s) for low resource PON applications. A number of methods were found unsuitable as they either involved multiple temperature incubations, were relatively expensive or required relatively large amounts target DNA for amplification. Among the methods examined, loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) were found to be the most suitable for PON applications as they are both single step methods that provide highly sensitive and reproducible amplifications. The speed of LAMP reactions was greatly enhanced, up to 76%, with the addition of loop primers while the presence of swarm primers and the sequestration of free magnesium ions with nucleotides also enhanced the amplification speed. In contrast, we were unable to enhance RPA's performance from the original published literature. While both RPA and LAMP have some drawbacks, either isothermal technology can reliably be used for on-site diagnostics with minimal equipment.

| S-EPMC7323990 | biostudies-literature

Isothermal amplification using a chemical heating device for point-of-care detection of HIV-1.

Project description:BACKGROUND: To date, the use of traditional nucleic acid amplification tests (NAAT) for detection of HIV-1 DNA or RNA has been restricted to laboratory settings due to time, equipment, and technical expertise requirements. The availability of a rapid NAAT with applicability for resource-limited or point-of-care (POC) settings would fill a great need in HIV diagnostics, allowing for timely diagnosis or confirmation of infection status, as well as facilitating the diagnosis of acute infection, screening and evaluation of infants born to HIV-infected mothers. Isothermal amplification methods, such as reverse-transcription, loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are ideal for POC settings, since they are typically quicker, easier to perform, and allow for integration into low-tech, portable heating devices. METHODOLOGY/SIGNIFICANT FINDINGS: In this study, we evaluated the HIV-1 RT-LAMP assay using portable, non-instrumented nucleic acid amplification (NINA) heating devices that generate heat from the exothermic reaction of calcium oxide and water. The NINA heating devices exhibited stable temperatures throughout the amplification reaction and consistent amplification results between three separate devices and a thermalcycler. The performance of the NINA heaters was validated using whole blood specimens from HIV-1 infected patients. CONCLUSION: The RT-LAMP isothermal amplification method used in conjunction with a chemical heating device provides a portable, rapid and robust NAAT platform that has the potential to facilitate HIV-1 testing in resource-limited settings and POC.

| S-EPMC3285652 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data