20 research outputs found
Establishment and application of cross-priming isothermal amplification coupled with lateral flow dipstick (CPA-LFD) for rapid and specific detection of red-spotted grouper nervous necrosis virus
Development and validation of a quantitative real-time polymerase chain assay for universal detection of the White Spot Syndrome Virus in marine crustaceans
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Rapid and visual Chlamydia trachomatis detection using loop-mediated isothermal amplification and hydroxynaphthol blue.
We developed an assay comprising crude DNA lysis by simple heat treatment coupled loop-mediated isothermal amplification with hydroxynaphthol blue for Chlamydia trachomatis detection (petty patent pending), and evaluated the developed assay for its feasibility as a one-step point-of-care detection on 284 endocervical swab specimens from clinically symptomatic C. trachomatis and healthy subjects. This assay is sensitive to 0·04 pg of ompA, specific with six primers targeting C. trachomatis ompA region, rapid (45 min total assay time), inexpensive (approx. 3 USD/reaction), does not require sophisticated instrumentation, and has comparable assay effectiveness (95% specificity, 90-100% sensitivity) to bacterial DNA isolation by a commercial kit coupled with polymerase chain reaction and gel electrophoresis (98-100% specificity, 87-100% sensitivity) based on the clinical samples test. The test result could be read by naked eye through the colour change from violet (negative) to sky blue (positive) for C. trachomatis-infected specimens. Further, this assay uses all safe chemical reagents and is hence safe to the users.Significance and impact of the studyChlamydia trachomatis is the major bacterial sexually transmitted disease worldwide. The clinical symptoms are broad, and chronic C. trachomatis infections could lead to blindness, ectopic pregnancy, sterility in males and females, and a higher risk of the development of cervical cancer. The result indicates the potential usefulness of our crude DNA lysis coupled loop-mediated isothermal amplification with hydroxynaphthol blue for a simple, rapid, specific, sensitive and cost-effective assay for C. trachomatis detection from suspected specimens. This assay offers an alternative in the clinical diagnosis of C. trachomatis in resource-limited health-care facilities and clinical laboratories in developing countries, and in field tests
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Rapid and visual Chlamydia trachomatis detection using loop-mediated isothermal amplification and hydroxynaphthol blue.
We developed an assay comprising crude DNA lysis by simple heat treatment coupled loop-mediated isothermal amplification with hydroxynaphthol blue for Chlamydia trachomatis detection (petty patent pending), and evaluated the developed assay for its feasibility as a one-step point-of-care detection on 284 endocervical swab specimens from clinically symptomatic C. trachomatis and healthy subjects. This assay is sensitive to 0·04 pg of ompA, specific with six primers targeting C. trachomatis ompA region, rapid (45 min total assay time), inexpensive (approx. 3 USD/reaction), does not require sophisticated instrumentation, and has comparable assay effectiveness (95% specificity, 90-100% sensitivity) to bacterial DNA isolation by a commercial kit coupled with polymerase chain reaction and gel electrophoresis (98-100% specificity, 87-100% sensitivity) based on the clinical samples test. The test result could be read by naked eye through the colour change from violet (negative) to sky blue (positive) for C. trachomatis-infected specimens. Further, this assay uses all safe chemical reagents and is hence safe to the users.Significance and impact of the studyChlamydia trachomatis is the major bacterial sexually transmitted disease worldwide. The clinical symptoms are broad, and chronic C. trachomatis infections could lead to blindness, ectopic pregnancy, sterility in males and females, and a higher risk of the development of cervical cancer. The result indicates the potential usefulness of our crude DNA lysis coupled loop-mediated isothermal amplification with hydroxynaphthol blue for a simple, rapid, specific, sensitive and cost-effective assay for C. trachomatis detection from suspected specimens. This assay offers an alternative in the clinical diagnosis of C. trachomatis in resource-limited health-care facilities and clinical laboratories in developing countries, and in field tests
Demonstration of a Very Inexpensive, Turbidimetric, Real-Time, RT-LAMP Detection Platform Using Shrimp Laem-Singh Virus (LSNV) as a Model
Sensitive Visual Detection of AHPND Bacteria Using Loop-Mediated Isothermal Amplification Combined with DNA-Functionalized Gold Nanoparticles as Probes
Use of reverse transcription loop-mediated isothermal amplification combined with lateral flow dipstick for an easy and rapid detection of Jembrana disease virus
Jembrana disease virus (JDV) is a viral pathogen that causes Jembrana disease in Bali cattle (Bos javanicus) with high mortality rate. An easy and rapid diagnostic method is essential for further control this disease. We used a reverse transcription loop-mediated isothermal amplification (RT-LAMP) combined with lateral flow dipstick (LFD), based on conserved tm subunit of Jembrana disease virus env gene. The RT-LAMP conditions were optimized by varying the concentration of MgSO(4), betaine, dNTP, and temperature as well as the time and duration of reaction. The primers sensitivity for JDV was confirmed. The method was able to detect env-tm gene dilution which contained 2 × 10(−15) g of template. Comparatively, the sensitivity of RT-LAMP/LFD was 100-fold more sensitive than reverse transcription-polymerase chain reaction. The primers specificity for JDV was also confirmed using positive and negative controls. This work also showed that virus detection could be done not only on total RNA extracted from blood but various organs could also be analyzed for the presence of JDV using RT-LAMP/LFD method. The whole process, including the LAMP reaction and the LFD hybridization step only lasts approximately 75 min. Results of analysis can be easily observed with naked eyes without addition of any chemical or further analysis. The combination of RT-LAMP with LFD makes the method a more suitable diagnostic tool in conditions where sophisticated and expensive equipments are not available for field investigations on Jembrana disease in Bali cattle