Development and evaluation of a 3D Printer-based DNA extraction method coupled to loop mediated isothermal amplification (LAMP) for point-of-care diagnosis of congenital Chagas disease in endemic regions

2020 ◽  
Author(s):  
Diana Patricia Wehrendt ◽  
Julio Alonso-Padilla ◽  
Bo Liu ◽  
Lizeth Rojas Panozo ◽  
Silvia Rivera Nina ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Chagas Disease ◽  
Extraction Method ◽  
Point Of Care ◽  
Isothermal Amplification ◽  
3D Printer ◽  
Loop Mediated Isothermal Amplification ◽  
Dna Extraction Method

scholarly journals Evaluation of different DNA extraction methods and loop-mediated isothermal amplification primers for the detection of Mycobacterium ulcerans in clinical specimens

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Anthony Ablordey ◽  
Evans Ahotor ◽  
Charles A. Narh ◽  
Sandra A. King ◽  
Isra Cruz ◽  
...  
Keyword(s):  
Dna Extraction ◽  
Extraction Method ◽  
Point Of Care ◽  
Buruli Ulcer ◽  
Extraction Methods ◽  
Isothermal Amplification ◽  
Mycobacterium Ulcerans ◽  
Loop Mediated Isothermal Amplification ◽  
Dna Extraction Method ◽  
Dna Extraction Methods

Abstract Background Early diagnosis and treatment of Buruli ulcer is critical in order to avoid the debilitating effects of the disease. In this regard, the development of new diagnostic and point of care tools is encouraged. The loop-mediated isothermal amplification for the detection of Mycobacterium ulcerans represents one of the new tools with a good potential of being developed into a point of care test. There is however the need to standardize the assays, reduce sample preparation times, improve the detection/visualization system and optimize them for high-throughput screening, adaptable to low resourced laboratories. Methods In this study, we assessed two DNA extraction protocols (modified Boom and EasyNAT methods), three previously published LAMP primer sets (BURULI, MU 2404 and BU-LAMP), and compared the sensitivity and specificity of LAMP assays on three DNA amplification platforms. Results Our results show that Buruli ulcer diagnosis using primers targeting IS2404 for the LAMP method is sensitive (73.75–91.49%), depending on the DNA extraction method used. Even though the modified Boom DNA extraction method provided the best results, its instrumentation requirement prevent it from being field applicable. The EasyNAT method on the other hand is simpler and may represent the best method for DNA extraction in less resourced settings. Conclusions For further work on the development and use of LAMP tests for Buruli diagnosis, it is recommended that the BURULI sets of primers be used, as these yielded the best results in terms of sensitivity (87.50–91.49%) and specificity (89.23–100%), depending on the DNA extraction methods used.

scholarly journals Ekstraksi DNA dari Daging Segar untuk Analisis dengan Metode Loop-Mediated Isothermal Amplification (LAMP)

2018 ◽  
Vol 4 (2) ◽  
Author(s):  
Rosy Hutami ◽  
Hanifah Bisyri ◽  
Sukarno Sukarno ◽  
Henny Nuraini ◽  
Raafqi Ranasasmita
Keyword(s):  
Dna Extraction ◽  
Extraction Method ◽  
Extraction Process ◽  
Extraction Methods ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification ◽  
Commercial Kits ◽  
Commercial Kit ◽  
Better Than

DNA extraction is needed in the analysis using the Loop-mediated isothermal amplification (LAMP) method because this method identifies nucleic acids. Some extraction methods that can be selected including commercial kits extraction method and phenol-chloroform extraction method. The purpose of this study was to obtain the best quality DNA extract between the two extraction methods. The DNA extraction process produced DNA concentrations between 31.06 - 410.18 ng / ml for the commercial kit DNA extract and 212.60 - 1502.30 ng / ml for the phenol-choroform DNA extract, while the purity of DNA were 1.82-2.02 for commercial kit DNA extract and 1.93-2.02 for phenol-chloroform DNA extract. The concentration and purity of extracts produced from both methods meet the requirements for molecular analysis. The purity and visualization results of commercial kit DNA extract are better than those produced from extraction from the phenol-chloroform method. DNA extract obtained from the commercial kit method was chosen to be used in the amplification stage of the method (LAMP).

Sensitivity detection of Abacavir in human through SNP detection of HLA-B*5701 allele

2016 ◽  
Vol 5 (08) ◽  
pp. 4754
Author(s):  
Tanushree Mitra* ◽  
Shivshankar Kumdale ◽  
Sameer Chowdhary ◽  
Amol D. Raut
Keyword(s):  
Blood Sample ◽  
Dna Extraction ◽  
Genomic Dna ◽  
Extraction Method ◽  
Snp Detection ◽  
Dna Extraction Method ◽  
Sensitivity Detection

The main objective of this study was to make sure whether randomly taken 12 samples were sensitive to abacavir. The genomic DNA from 12 blood sample were extracted by phenol chloroform DNA extraction method, extracted genomic DNA were amplified and sequenced, thereafter SNPs were detected. Every sample had shown the presence of normal base at SNP position. This study indicated, those randomly taken 12 patients were sensitive to abacavir, so they can consume abacavir if they get infected with HIV.

scholarly journals Essential properties and pitfalls of colorimetric Reverse Transcription Loop-mediated Isothermal Amplification as a point-of-care test for SARS-CoV-2 diagnosis

2021 ◽  
Vol 27 (1) ◽  
Author(s):  
Bruna de Oliveira Coelho ◽  
Heloisa Bruna Soligo Sanchuki ◽  
Dalila Luciola Zanette ◽  
Jeanine Marie Nardin ◽  
Hugo Manuel Paz Morales ◽  
...  
Keyword(s):  
Viral Load ◽  
Internal Control ◽  
Reverse Transcription ◽  
Color Change ◽  
Point Of Care ◽  
Colorimetric Detection ◽  
False Negative ◽  
Isothermal Amplification ◽  
Clinical Samples ◽  
Loop Mediated Isothermal Amplification

Abstract Background SARS-CoV-2 Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP) colorimetric detection is a sensitive and specific point-of-care molecular biology technique used to detect the virus in only 30 min. In this manuscript we have described a few nuances of the technique still not properly described in the literature: the presence of three colors clusters; the correlation of the viral load with the color change; and the importance of using an internal control to avoid false-negative results. Methods To achieve these findings, we performed colorimetric RT-LAMP assays of 466 SARS-CoV-2 RT-qPCR validated clinical samples, with color quantification measured at 434 nm and 560 nm. Results First we determinate a sensitivity of 93.8% and specificity of 90.4%. In addition to the pink (negative) and yellow (positive) produced colors, we report for the first time the presence of an orange color cluster that may lead to wrong diagnosis. We also demonstrated using RT-qPCR and RT-LAMP that low viral loads are related to Ct values > 30, resulting in orange colors. We also demonstrated that the diagnosis of COVID-19 by colorimetric RT-LAMP is efficient until the fifth symptoms day when the viral load is still relatively high. Conclusion This study reports properties and indications for colorimetric RT-LAMP as point-of-care for SARS-CoV-2 diagnostic, reducing false results, interpretations and optimizing molecular diagnostics tests application.

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