Digital microfluidics with distance-based detection - a new approach for nucleic acid diagnostics

被引:12
|
作者
Ho, Man [1 ,2 ]
Sathishkumar, N. [1 ,2 ]
Sklavounos, Alexandros A. [1 ,2 ]
Sun, Jianxian [1 ]
Yang, Ivy [3 ]
Nichols, Kevin P. [4 ]
Wheeler, Aaron R. [1 ,2 ,5 ]
机构
[1] Univ Toronto, Dept Chem, 80 St George St, Toronto, ON M5S 3H6, Canada
[2] Univ Toronto, Donnelly Ctr Cellular & Biomol Res, 160 Coll St, Toronto, ON M5S 3E1, Canada
[3] Univ Toronto, Dept Chem Engn & Appl Chem, 200 Coll St, Toronto, ON M5S 3E5, Canada
[4] Global Hlth Labs, Bellevue, WA 98007 USA
[5] Univ Toronto, Inst Biomat & Biomed Engn, 164 Coll St, Toronto, ON M5S 3G9, Canada
基金
加拿大自然科学与工程研究理事会;
关键词
POLYMERASE-CHAIN-REACTION; AMPLIFICATION; PLATFORM; QUANTIFICATION; DEVICE; SYSTEM; DNA;
D O I
10.1039/d3lc00683b
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
There is great enthusiasm for using loop-mediated isothermal amplification (LAMP) in point-of-care nucleic acid amplification tests (POC NAATs), as an alternative to PCR. While isothermal amplification techniques like LAMP eliminate the need for rapid temperature cycling in a portable format, these systems are still plagued by requirements for dedicated optical detection apparatus for analysis and manual off-chip sample processing. Here, we developed a new microfluidic system for LAMP-based POC NAATs to address these limitations. The new system combines digital microfluidics (DMF) with distance-based detection (DBD) for direct signal readout. This is the first report of the use of (i) LAMP or (ii) DMF with DBD - thus, we describe a number of characterization steps taken to determine optimal combinations of reagents, materials, and processes for reliable operation. For example, DBD was found to be quite sensitive to background signals from low molecular weight LAMP products; thus, a Capto (TM) adhere bead-based clean-up procedure was developed to isolate the desirable high-molecular-weight products for analysis. The new method was validated by application to detection of SARS-CoV-2 in saliva. The method was able to distinguish between saliva containing no virus, saliva containing a low viral load (104 genome copies per mL), and saliva containing a high viral load (108 copies per mL), all in an automated system that does not require detection apparatus for analysis. We propose that the combination of DMF with distance-based detection may be a powerful one for implementing a variety of POC NAATs or for other applications in the future. We introduce the integration of digital microfluidics (DMF) with distance-based detection (DBD). This platform allows for miniaturized diagnosis of infectious diseases in a format that is easy to operate with a detection scheme that can be read by eye.
引用
收藏
页码:63 / 73
页数:11
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