1. Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP
- Author
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Friedrich Zitz, Hans Nitschko, Mohammed A. Bakheit, Karl Stock, Florian Hausladen, Dominik Drossart, Nils Paust, Giampiero Porro, Oliver Strohmeier, Roland Zengerle, Felix von Stetten, Nadine Borst, Hannes Hofer, Silvina Stinco, Mark Keller, Holger Wurm, Lisa Becherer, Dominik Huber, Simon Wadle, Sieghard Frischmann, Hanna Kuhn, Jacob Friedrich Hess, and Tobias Hutzenlaub
- Subjects
T-Lymphozyt ,Viral nucleic acid ,lcsh:Mechanical engineering and machinery ,Cell ,human T-cell lymphotropic virus 1 ,Loop-mediated isothermal amplification ,digital loop-mediated isothermal amplification ,02 engineering and technology ,01 natural sciences ,Article ,DDC 570 / Life sciences ,Mediator ,ddc:570 ,proviral load measurements ,medicine ,lcsh:TJ1-1570 ,ddc:610 ,Electrical and Electronic Engineering ,Whole blood ,Point of care ,Human T-lymphotropic virus 1 ,Chemistry ,Mechanical Engineering ,010401 analytical chemistry ,021001 nanoscience & nanotechnology ,Molecular biology ,Reverse transcriptase ,0104 chemical sciences ,Point-of-care testing ,medicine.anatomical_structure ,Control and Systems Engineering ,centrifugal microfluidics ,Nucleic acid ,0210 nano-technology ,DDC 610 / Medicine & health - Abstract
This paper presents a universal point-of-care system for fully automated quantification of human T-cell lymphotropic virus type 1 (HTLV-1) proviral load, including genomic RNA, based on digital reverse RNA transcription and c-DNA amplification by MD LAMP (mediator displacement loop-mediated isothermal amplification). A disposable microfluidic LabDisk with pre-stored reagents performs automated nucleic acid extraction, reaction setup, emulsification, reverse transcription, digital DNA amplification, and quantitative fluorogenic endpoint detection with universal reporter molecules. Automated nucleic acid extraction from a suspension of HTLV-1-infected CD4+ T-lymphocytes (MT-2 cells) yielded 8 ± 7 viral nucleic acid copies per MT-2 cell, very similar to the manual reference extraction (7 ± 2 nucleic acid copies). Fully automated sample processing from whole blood spiked with MT-2 cells showed a comparable result of 7 ± 3 copies per MT-2 cell after a run time of two hours and 10 min.
- Published
- 2021
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