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Development of an on-disc isothermal in vitro amplification and detection of bacterial RNA

Authors: 

Des Brennan, Helena Coughlan, Eoin Clancy, Nikolay Dimov, Thomas Barry, David Kinahane, Jens Ducréee, Terry Smith, Paul Galvin

Publication Type: 
Refereed Original Article
Abstract: 
We present a centrifugal microfluidic “Lab-on-a-Disc” (LoaD) system capable of implementing nucleic acid in vitro amplification using non-contact heating and fluorescence detection. The system functionality is verified by implementing a Nucleic Acid Sequence Based Amplification (NASBA) reaction, targeting the tmRNA transcript of Haemophilus influenzae. The NASBA assay incorporates fluorescent molecular beacon probes reporting target tmRNA amplification for endpoint detection. The system implements non-contact IR heating to heat the NASBA reaction to the required target temperatures during denaturation and amplification steps. The LoaD control system facilitates spin speed and chamber positioning for heating and fluorescence detection. The LoaD alignment system uses magnetic fields to locate and lock the chamber in the required position (heating or detection). The NASBA assay was implemented on the system using Haemophilus influenzae tmRNA over the range 102–104 cell equivalent (CE) units. For comparison, identical qNASBA assays were implemented on a Roche LightCycler 2.0 over this concentration range.
Digital Object Identifer (DOI): 
10.1016/j.snb.2016.08.018
Publication Status: 
Published
Date Accepted for Publication: 
Tuesday, 2 August, 2016
Publication Date: 
03/08/2016
Journal: 
Sensors and Actuators B: Chemical
Research Group: 
Institution: 
Tyndall National Institute
Open access repository: 
No