Automated Microfluidic Nucleic Acid Detection Platform-Integrated RPA-T7-Cas13a for Pathogen Diagnosis

Li，Zheng1; Hua，Liyan2; Xie，Liming2; Wang，Dou1; Jiang，Xingyu1

2023-05-02

10.1021/acs.analchem.3c00242

Analytical Chemistry   影响因子和分区

0003-2700

1520-6882

There is a growing urgent need for point-of-care testing (POCT) devices that integrate sample pretreatment and nucleic acid detection in a rapid, economical, and non-labor-intensive way. Here, we have developed an automated, portable nucleic acid detection system employing microfluidic chips integrating rotary valve-assisted sample pretreatment and recombinase polymerase amplification (RPA)-T7-Cas13a into one-step nucleic acid detection. The RPA and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas13a were integrated into a single-chamber reaction. As a validation model, we used this method to detect Group B streptococci (GBS) DNA and achieved a detection sensitivity of 8 copies/reaction, which is 6 times more sensitive than gold-standard polymerase chain reactions (PCRs). Dual specific recognition of RPA with CRISPR/Cas13a makes our method ultraspecific, with correct detection of Group B streptococci from 8 kinds of pathogenic bacteria. For the 16 positive and 24 negative clinical GBS samples, our assay achieved 100% accuracy compared to the PCR technique. The whole procedure can be automatically completed within 30 min, providing a more robust, sensitive, and accurate molecular diagnostic tool for POCT.

SCI ; EI

英语

NI论文

第一 ; 通讯

National Natural Science Foundation of China[22234004];

Chemistry

Chemistry, Analytical

WOS:000979377200001

AMER CHEMICAL SOC

20231814040851

Bacteria ; Biomolecules ; Microfluidic chips ; Microfluidics

Biology:461.9 ; Microfluidics:632.5.1 ; Biochemistry:801.2

CHEMISTRY

2-s2.0-85154018633

Scopus

1.Shenzhen Key Laboratory of Smart Healthcare Engineering,Guangdong Provincial Key Laboratory of Advanced Biomaterials,Department of Biomedical Engineering,Southern University of Science and Technology,Shenzhen,No. 1088, Xueyuan Road, Guangdong,518055,China
2.Shaoguan Maternal and Child Health Hospital,Shaoguan,No. 3, Dongdi South Road, Guangdong,512026,China

Li，Zheng,Hua，Liyan,Xie，Liming,et al. Automated Microfluidic Nucleic Acid Detection Platform-Integrated RPA-T7-Cas13a for Pathogen Diagnosis[J]. Analytical Chemistry,2023,95(17):6940-6947.

Li，Zheng,Hua，Liyan,Xie，Liming,Wang，Dou,&Jiang，Xingyu.(2023).Automated Microfluidic Nucleic Acid Detection Platform-Integrated RPA-T7-Cas13a for Pathogen Diagnosis.Analytical Chemistry,95(17),6940-6947.

Li，Zheng,et al."Automated Microfluidic Nucleic Acid Detection Platform-Integrated RPA-T7-Cas13a for Pathogen Diagnosis".Analytical Chemistry 95.17(2023):6940-6947.