Dynamic Aqueous Multiphase Reaction System for One-Pot CRISPR-Cas12a-Based Ultrasensitive and Quantitative Molecular Diagnosis
Document Type
Article
Publication Date
5-10-2020
Publication Title
Analytical Chemistry
Volume
92
Issue
12
First page number:
8561
Last page number:
8568
Abstract
Recently, CRISPR-Cas technology has opened a new era of nucleic acid-based molecular diagnostics. However, current CRISPR-Cas-based nucleic acid biosensing has a lack of the quantitative detection ability and typically requires separate manual operations. Herein, we reported a dynamic aqueous multiphase reaction (DAMR) system for simple, sensitive and quantitative one-pot CRISPR-Cas12a based molecular diagnosis by taking advantage of density difference of sucrose concentration. In the DAMR system, recombinase polymerase amplification (RPA) and CRISPR-Cas12a derived fluorescent detection occurred in spatially separated but connected aqueous phases. Our DAMR system was utilized to quantitatively detect human papillomavirus (HPV) 16 and 18 DNAs with sensitivities of 10 and 100 copies within less than 1 h. Multiplex detection of HPV16/18 in clinical human swab samples were successfully achieved in the DAMR system using 3D-printed microfluidic device. Furthermore, we demonstrated that target DNA in real human plasma samples can be directly amplified and detected in the DAMR system without complicated sample pretreatment. As demonstrated, the DAMR system has shown great potential for development of next-generation point-of-care molecular diagnostics.
Disciplines
Chemistry | Physical Sciences and Mathematics
Language
English
Repository Citation
Yin, K.,
Ding, X.,
Li, Z.,
Zhao, H.,
Cooper, K.,
Liu, C.
(2020).
Dynamic Aqueous Multiphase Reaction System for One-Pot CRISPR-Cas12a-Based Ultrasensitive and Quantitative Molecular Diagnosis.
Analytical Chemistry, 92(12),
8561-8568.
http://dx.doi.org/10.1021/acs.analchem.0c01459