Clustered Regularly Interspaced short palindromic repeats-Based Microfluidic System in Infectious Diseases Diagnosis: Current Status, Challenges, and Perspectives

Abstract

Mitigating the spread of global infectious diseases requires rapid and accurate diagnostic tools. Conventional diagnostic techniques for infectious diseases typically require sophisticated equipment and are time consuming. Emerging clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated proteins (Cas) detection systems have shown remarkable potential as next-generation diagnostic tools to achieve rapid, sensitive, specific, and field-deployable diagnoses of infectious diseases, based on state-of-the-art microfluidic platforms. Therefore, a review of recent advances in CRISPR-based microfluidic systems for infectious diseases diagnosis is urgently required. This review highlights the mechanisms of CRISPR/Cas biosensing and cutting-edge microfluidic devices including paper, digital, and integrated wearable platforms. Strategies to simplify sample pretreatment, improve diagnostic performance, and achieve integrated detection are discussed. Current challenges and future perspectives contributing to the development of more effective CRISPR-based microfluidic diagnostic systems are also proposed.

Keywords: clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats-associated proteins biosensing mechanisms; infectious diseases diagnosis; integrated detection; microfluidic platforms.

Figure 1

Overview of CRISPR‐based microfluidic system in infectious diseases diagnosis.

Figure 2

Workflow of sample preparation in CRISPR‐based diagnosis.

Figure 3

CRISPR‐based biosensing system. Three CRISPR‐based cleavage mechanisms (Cas9, Cas13, and Cas12) and three fundamental CRISPR‐based biosensing strategies (NASBACC, SHERLOCK, and DETECTR) are summarized.

Figure 4

Amplification‐free CRISPR‐based biosensing. A) Multiple crRNAs and digital strategy. B) Signaling cascade amplification. C) Nanoparticle‐assisted strategy.

Figure 5

Polymer‐based microfluidic devices. A) The illustration of CRISPR‐based surface‐enhanced Raman spectroscopy‐active nanoarray. Reproduced with permission.^{[ 111 ]} Copyright 2021, American Chemical Society. B) The illustration of CASMEAN. Reproduced with permission.^{[ 121 ]} Copyright 2020, American Chemical Society. C) The description of the IMPACT chip. The chip comprises a PDMS micropillar with surface treatment and ssDNA probe binding to achieve CRISPR detection.

Figure 6

Paper‐based microfluidic devices. A) Workflow of advanced CASLFA method and construction of the lateral‐flow chip. B) Device constitution and workflow of hybrid paper‐based CRISPR chambers for multiplex gene diagnosis. C) Device constitution and workflow of µReaCH‐PAD.

Figure 7

Electronic CRISPR microfluidic devices. A) Illustration of CRISPR‐biosensor “X” and the workflow on this electrode. B) Illustration of “on–off” signal‐switchable sensor and the workflow on this chamber. C) Illustration of CRISPR Cas13a‐gFET. Reproduced with permission.^{[ 150 ]} Copyright 2022, Wiley‐VCH GmbH.

Figure 8

Digital microfluidic devices. A) Schematic of the ultralocalized Cas13a assay on microfluidic droplet chip. B) Illustration of deCOViD. C) Schematic and comparison of CARMEN v.1 and mCARMEN workflow.

Figure 9

Wearable microfluidic devices. A) The facemask‐integrated SARS‐CoV‐2 wearable device and the schematic of the SARS‐CoV‐2 sensor components. Reproduced with permission.^{[ 15 ]} Copyright 2021, Springer Nature Limited. B) CRISPR‐based multi‐sensor. Reproduced with permission.^{[ 15 ]} Copyright 2021, Springer Nature Limited. C) CRISPR microneedles platform.

Figure 10

Future perspectives in CRISPR‐based microfluidic technologies.