Browse
Submit Data
Databases
API
Help

Dataset Information

17 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Loop-Mediated Isothermal Amplification Assay for Detecting Tumor Markers and Human Papillomavirus: Accuracy and Supplemental Diagnostic Value to Endovaginal MRI in Cervical Cancer

ABSTRACT:

Objective

To establish the sensitivity and specificity of a human papillomavirus (HPV) and tumor marker DNA/mRNA assay for detecting cervical cancer that is transferrable to a Lab-on-a-chip platform and determine its diagnostic benefit in early stage disease when used in conjunction with high-resolution endovaginal magnetic resonance imaging (MRI).

Methods

Forty-one patients (27 with Stage1 cervical cancer [Group1] and 14 non-cancer HPV negative controls [Group2]) had DNA and RNA extracted from cervical cytology swab samples. HPV16, HPV18, hTERT, TERC/GAPDH and MYC/GAPDH concentration was established using a loop mediated isothermal amplification (LAMP) assay. Thresholds for tumor marker detection for Group1 were set from Group2 analysis (any hTERT, TERC/GAPDH 3.12, MYC/GAPDH 0.155). Group 1 participants underwent endovaginal MRI. Sensitivity and specificity for cancer detection by LAMP and MRI individually and combined was documented by comparison to pathology.

Results

Sensitivity and specificity for cancer detection was 68.8% and 77.8% if any tumor marker was positive regardless of HPV status (scenario1), and 93.8% and 55.8% if tumor marker or HPV were positive (scenario 2). Adding endovaginal MRI improved specificity to 88.9% in scenario 1 (sensitivity 68.8%) and to 77.8%% in scenario2 (sensitivity 93.8%).

Conclusion

Specificity for cervical cancer detection using a LAMP assay is superior with tumor markers; low sensitivity is improved by HPV detection. Accuracy for early stage cervical cancer detection is optimal using a spatially multiplexed tumor marker/HPV LAMP assay together with endovaginal MRI.

SUBMITTER: Wormald B

PROVIDER: S-EPMC8591099 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

A novel loop-mediated isothermal amplification-based test for detecting Neospora caninum DNA.

Project description:Neospora caninum is a cyst-forming, coccidian parasite which is known to cause neurological disorders in dogs and abortion and neonatal mortality in cows and other livestock. This study reports the development of a loop-mediated isothermal amplification (LAMP) assay based on the Neospora caninum Nc-5 gene and compares its efficacy for detecting DNA to that of a semi-nested PCR test.Six primers were designed based on the Nc-5 repeat region of N. caninum. Specific LAMP primers led to successful amplification of N. caninum DNA at 63 °C in 30 min. The LAMP assay was highly specific (i.e. it did not reveal cross-reactivity with other parasite species) and had a low N. caninum plasmid DNA limit of detection (1 fg), which is ten times higher than that for the semi-nested PCR. LAMP applicability was evaluated using a set of naturally-infected samples (59 from canine faeces and five from bovine abortions). Thirty-nine percent (25/64) of the naturally-infected samples were positive for N. caninum DNA by LAMP and 36% (23/64) by semi-nested PCR. However, the LAMP assay is much faster to perform than semi-nested PCR and provides results in 30 min.The optimized reaction conditions described in this study resulted in a sensitive, specific and rapid technique for detecting N. caninum DNA. Considering the advantages of LAMP for detecting N. caninum DNA, further assays aimed at testing its usefulness on a wider range of field samples are recommended.

| S-EPMC5707868 | biostudies-literature

A quantitative loop-mediated isothermal amplification assay for detecting a novel goose astrovirus.

Project description:In November 2017, a severe infectious disease that devastated the major goose-producing regions in China was found to be caused by a novel goose astrovirus (N-AstV). The objective of this study was to develop a quantitative loop-mediated isothermal amplification (qLAMP) assay for the rapid diagnosis of N-AstV characterized with gout, hemorrhage, and swellings of the kidneys. A set of 4 specific primers, 2 inner and 2 outer primers, targeting the ORF1a gene of N-AstV were designed for the assay which could be completed within 60 min at 65°C in a water bath or on a real-time PCR instrument for quantitative analysis. The qLAMP assay showed a high sensitivity with a detection limit of 1 × 101 copies of the target DNA/μL. There were no cross-reactions with other viruses, and the reproducibility of the assay was confirmed in intrasensitivity and intersensitivity assay tests with variability ranging from 0.61 to 2.21%. The results indicated that the qLAMP assay for N-AstV was a simple, accurate, rapid, sensitive, and specific, especially useful for field detection.

| S-EPMC7705033 | biostudies-literature

Molecular Markers for Detecting <i>Schistosoma</i> Species by Loop-Mediated Isothermal Amplification.

Project description:Schistosomiasis is considered a neglected parasitic disease. Around 280,000 people die from it annually, and more than 779 million people are at risk of getting infected. The schistosome species which infect human beings are Schistosoma mansoni, Schistosoma haematobium, Schistosoma intercalatum, Schistosoma japonicum, Schistosoma guineensis, and Schistosoma mekongi. This disease is also of veterinary significance; the most important species being Schistosoma bovis since it causes the disease in around 160 million livestock in Africa and Asia. This work was aimed at designing and developing a genus-specific loop-mediated isothermal amplification (LAMP) method for detecting the most important schistosome species affecting humans and for the species-specific detection of S. bovis. Bioinformatics tools were used for primer design, and the LAMP method was standardised for detecting the ITS-1 region from S. intercalatum, S. haematobium, S. mansoni, S. japonicum, and S. bovis DNA (generic test) and the NADH 1 gene for specifically detecting S. bovis (at different DNA concentrations). Detection limits achieved were 1 pg DNA for S. mansoni, 0.1 pg for S. haematobium, 1 pg for S. intercalatum, and 10 pg for S. bovis. No amplification for S. japonicum DNA was obtained. The LAMP designed for the amplification of S. bovis NADH-1 worked specifically for this species, and no other DNA from other schistosome species included in the study was amplified. Two highly sensitive LAMP methods for detecting different Schistosoma species important for human and veterinary health were standardised. These methods could be very useful for the diagnosis and surveillance of schistosome infections.

| S-EPMC7396120 | biostudies-literature

Accuracy of loop-mediated isothermal amplification for diagnosis of human leptospirosis in Thailand.

Project description:There is a lack of diagnostic tests for leptospirosis in technology-restricted settings. We developed loop-mediated isothermal amplification (LAMP) specific for the 16S ribosomal RNA gene (rrs) of pathogenic and intermediate group Leptospira species. The lower limit of detection was 10 genomic equivalents/reaction, and analytical specificity was high; we observed positive reactions for pathogenic/intermediate groups and negative reactions for non-pathogenic Leptospira species and other bacterial species. We evaluated this assay in Thailand by using a case-control study of 133 patients with laboratory-proven leptospirosis and 133 patients with other febrile illnesses. Using admission blood, we found that the rrs LAMP showed positive results in 58 of 133 cases (diagnostic sensitivity = 43.6, 95% confidence interval [CI] = 35.0-52.5) and in 22 of 133 controls (diagnostic specificity = 83.5, 95% CI = 76.0-89.3). Sensitivity was high for 39 patients who were culture positive for Leptospira spp. (84.6, 95% CI = 69.5-94.1). The rrs LAMP can provide an admission diagnosis in approximately half of patients with leptospirosis, but its clinical utility is reduced by a lower specificity.

| S-EPMC3062458 | biostudies-literature

Development of loop-mediated isothermal amplification methods for detecting Taylorella equigenitalis and Taylorella asinigenitalis.

Project description:Taylorella equigenitalis is a causative bacterium of contagious equine metritis (CEM), and Taylorella asinigenitalis is species belonging to genus Taylorella. The authors developed two loop-mediated isothermal amplification (LAMP) methods, Te-LAMP and Ta-LAMP, for detecting T. equigenitalis and T. asinigenitalis, respectively. Using experimentally spiked samples, Te-LAMP was as sensitive as a published semi-nested PCR method, and Ta-LAMP was more sensitive than conventional PCR. Multiplex LAMP worked well without nonspecific reactions, and the analytical sensitivities of multiplex LAMP in the spiked samples were almost equivalent to those of Te-LAMP and Ta-LAMP. Therefore, the LAMP methods are considered useful tools to detect T. equigenitalis and/or T. asinigenitalis, and preventive measures will be rapidly implemented if the occurrence of CEM is confirmed by the LAMP methods.

| S-EPMC4379330 | biostudies-literature

One-Pot Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP) for Detecting MERS-CoV.

Project description:Due to the limitation of rapid development of specific antiviral drug or vaccine for novel emerging viruses, an accurate and rapid diagnosis is a key to manage the virus spread. We developed an efficient and rapid method with high specificity for the Middle East Respiratory Syndrome coronavirus (MERS-CoV), based on one-pot reverse transcription loop-mediated isothermal amplification (one-pot RT-LAMP). A set of six LAMP primers [F3, B3, FIP, BIP, LF (Loop-F), and LB (Loop-B)] were designed using the sequence of nucleocapsid (N) gene with optimized RT-LAMP enzyme conditions: 100 U M-MLV RTase and 4 U Bst polymerase, implying that the reaction was able to detect four infectious viral genome copies of MERS-CoV within a 60 min reaction time period. Significantly, EvaGreen dye has better signal read-out properties in one-pot RT-LAMP reaction and is more compatible with DNA polymerase than SYBR green I. Isothermally amplified specific N genes were further evaluated using field-deployable microchamber devices, leading to the specific identification of as few as 0.4 infectious viral genome copies, with no cross-reaction to the other acute respiratory disease viruses, including influenza type A (H1N1 and H3N2), type B, human coronavirus 229E, and human metapneumovirus. This sensitive, specific and feasible method provides a large-scale technical support in emergencies, and is also applied as a sample-to-detection module in Point of Care Testing devices.

| S-EPMC5220095 | biostudies-literature

Loop-mediated isothermal amplification linked a nanoparticles-based biosensor for detecting Epstein-Barr virus.

Project description:Epstein-Barr virus (EBV) is a ubiquitous gamma herpesvirus that maintains a lifelong latent association with B lymphocytes. Here, a rapid and reliable diagnosis platform for detecting EBV infection, employing loop-mediated isothermal amplification (LAMP) combined with a gold nanoparticles-based lateral flow biosensors (AuNPs-LFB) (termed LAMP Amplification Mediated AuNPs-LFB Detection, LAMAD), was developed in the current study. A set of specific LAMP primers targeting the Epstein-Barr nuclear antigen (EBNA) leader protein (EBNA-LP) gene was designed and synthesized. Subsequently, these templates extracted from various pathogens and whole blood samples were used to optimize and evaluate the EBV-LAMAD assay. As a result, the limit of detection (LoD) of the EBV-LAMAD assay was 45 copies/reaction. The EBV-LAMAD assay can detect all representative EBV pathogens used in the study, and of note, no cross-reactions were observed with other non-EBV organisms. Moreover, the whole workflow of the EBV-LAMAD assay can be completed within 70 min, including rapid EBV template preparation, EBV-LAMP amplification, and AuNPs-LFB-mediated detection. Taken together, the EBV-LAMAD assay targeting the EBNA-LP gene is a rapid, simplified, sensitive, reliable, and easy-to-use detection protocol that can be used as a competitive potential diagnostic/screening tool for EBV infection in clinical settings, especially in basic laboratories in resource-limited regions. KEY POINTS: • A novel, simplified, and easy-to-use AuNPs-LFB biosensor was designed and prepared. • LAMP combined with an AuNPs-LFB targeting the novel EBNA-LP gene was established. • EBV-LAMAD is a rapid, sensitive, and reliable detection protocol for EBV infection.

| S-EPMC10784390 | biostudies-literature

Development of a toxR-based loop-mediated isothermal amplification assay for detecting Vibrio parahaemolyticus.

Project description:BackgroundVibrio parahaemolyticus is a leading cause of seafood-related bacterial gastroenteritis and outbreaks worldwide. Sensitive and specific detection methods are needed to better control V. parahaemolyticus infections. This study aimed at developing a highly specific and sensitive loop-mediated isothermal amplification (LAMP) assay for detecting V. parahaemolyticus in oysters. A set of five LAMP primers, two outer, two inner, and one loop were designed based on the published V. parahaemolyticus toxR sequence. Specificity of the assay was evaluated using a panel of 36 V. parahaemolyticus and 39 other strains. The assay sensitivity was determined using serial dilutions of V. parahaemolyticus ATCC 27969 culture ranging from 10(8) CFU/ml to extinction. The assay was also tested in experimentally inoculated oyster samples.ResultsThe toxR-based LAMP assay was able to specifically detect all of the 36 V. parahaemolyticus strains without amplification from 39 other strains. The detection limit was 47-470 cells per reaction in pure culture, up to 100-fold more sensitive than that of toxR-PCR. When applied in spiked oysters, the assay was able to detect 1.1 x 10(5) V. parahaemolyticus cells per gram of oyster without enrichment, up to 100-fold more sensitive than that of toxR-PCR. Standard curves generated for detecting V. parahaemolyticus in both pure culture and spiked oyster samples showed good linear relationship between cell numbers and the fluorescence or turbidity signals.ConclusionsThe toxR-based LAMP assay developed in this study was sensitive, specific, and quantitative, holding great potential for future field detection of V. parahaemolyticus in raw oysters.

| S-EPMC2838873 | biostudies-literature

RNase H-dependent amplification improves the accuracy of rolling circle amplification combined with loop-mediated isothermal amplification (RCA-LAMP).

Project description:The hybrid method upon combining rolling circle amplification and loop-mediated isothermal amplification (RCA-LAMP) was developed to quantify very small amount of different type of RNAs, such as miRNAs. RCA-LAMP can help detect short sequences through padlock probe (PLP) circularization and exhibit powerful DNA amplification. However, one of the factors that determines the detection limit of RCA-LAMP is non-specific amplification. In this study, we improved the accuracy of RCA-LAMP through applying RNase H-dependent PCR (rhPCR) technology. In this method, the non-specific amplification was suppressed by using the rh primer, which is designed through blocking the modification at the 3'end to stop DNA polymerase reaction and replacing the 6th DNA molecule from the end with RNA using RNase H2 enzyme. Traditional RCA-LAMP amplified the non-specific amplicons from linear PLP without a targeting reaction, while RCA-LAMP with rh primer and RNase H2 suppressed the non-specific amplification. Conversely, we identified the risk posed upon conducting PLP cyclization reaction using Splint R ligase in the RNA-targeting step that occurred even in the RNA-negative condition, which is another factor determining the detection limit of RCA-LAMP. Therefore, this study contributes in improving the accuracy of RNA quantification using RCA-LAMP.

| S-EPMC8327969 | biostudies-literature

Detecting wMel Wolbachia in field-collected Aedes aegypti mosquitoes using loop-mediated isothermal amplification (LAMP).

Project description:BackgroundThe World Mosquito Program uses Wolbachia pipientis for the biocontrol of arboviruses transmitted by Aedes aegypti mosquitoes. Diagnostic testing for Wolbachia in laboratory colonies and in field-caught mosquito populations has typically employed PCR. New, simpler methods to diagnose Wolbachia infection in mosquitoes are required for large-scale operational use.MethodsField-collected Ae. aegypti mosquitoes from North Queensland were tested using primers designed to detect the Wolbachia wsp gene, specific to the strain wMel. The results were analysed by colour change in the reaction mix. Furthermore, to confirm the efficiency of the LAMP assay, the results were compared to the gold-standard qPCR test.ResultsA novel loop-mediated isothermal amplification (LAMP) colorimetric test for the wMel strain of Wolbachia was designed, developed and validated for use in a high-throughput setting. Against the standard qPCR test, the analytical sensitivity, specificity and diagnostic metrics were: sensitivity (99.6%), specificity (92.2%), positive predictive value (97.08%) and negative predictive value (99.30%).ConclusionsWe describe an alternative, novel and high-throughput method for diagnosing wMel Wolbachia infections in mosquitoes. This assay should support Wolbachia surveillance in both laboratory and field populations of Ae. aegypti.

| S-EPMC6694616 | 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