CRISPR-powered electrochemical microfluidic multiplexed biosensor for target amplification-free miRNA diagnostics

Bruch, Richard C.; Johnston, Midori; Kling, André; Mattmüller, Thorsten; Baaske, Julia; Partel, Stefan; Madlener, Sibylle; Weber, Wilfried; Urban, G.; Dincer, Can

doi:10.1016/j.bios.2020.112887

Cited by 136 publications

(98 citation statements)

References 18 publications

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“…As an improved CRISPR detection system, CECRID can be applied in more broad scenarios of nucleic acid detection. In addition to fluorescence or lateral flow strip readout, CECRID, as an enhanced reaction system, could be combined with other advanced or integrated detection systems such as electrochemical microfluidic biosensor to develop more practical and powerful diagnostic tools ( Bruch et al 2019 , 2021 ). Taken together, our work not only uncovers several key parameters affecting the strength of nucleic acid detection, but also paves the way for developing robust reagent formula or test kit towards practical POCT application of CRISPR-based detection technology.…”

Section: Discussionmentioning

confidence: 99%

A chemical-enhanced system for CRISPR-Based nucleic acid detection

Zhao

et al. 2021

Biosensors and Bioelectronics

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The CRISPR-based nucleic acid detection systems have shown great potential for point-of-care testing of viral pathogens, especially in the context of COVID-19 pandemic. Here we optimize several key parameters of reaction chemistry and develop a Chemical Enhanced CRISPR Detection system for nucleic acid (termed CECRID). For the Cas12a/Cas13a-based signal detection phase, we determine buffer conditions and substrate range for optimal detection performance, and reveal a crucial role of bovine serum albumin in enhancing trans-cleavage activity of Cas12a/Cas13a effectors. By comparing several chemical additives, we find that addition of L-proline can secure or enhance Cas12a/Cas13a detection capability. For isothermal amplification phase with typical LAMP and RPA methods, inclusion of L-proline can also enhance specific target amplification as determined by CRISPR detection. Using SARS-CoV-2 pseudovirus, we demonstrate CECRID has enhanced detection sensitivity over chemical additive-null method with either fluorescence or lateral flow strip readout. Thus, CECRID provides an improved detection power and system robustness, and helps to develop enhanced reagent formula or test kit towards practical application of CRISPR-based diagnostics.

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Section: Discussionmentioning

confidence: 99%

A chemical-enhanced system for CRISPR-Based nucleic acid detection

Zhao

et al. 2021

Biosensors and Bioelectronics

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“…Combining with sensor array or microfluidics, it is potential for CRISPR-Cas to win wide application in multiplex detection ( Li et al, 2019c ). Bruch et al designed a CRISPR based electrochemical microfluidic multiplexed biosensor through fabrication of multiple sensing channels ( Bruch et al, 2021 ). For quantitative testing, CRISPR-Cas based assay is difficult to quantify the targets with high concentration, which is easy to reach the signal plateau because of the high sensitivity of the system and the limited reporter molecules ( Li et al, 2019a ).…”

Section: Challenges and Perspectivesmentioning

confidence: 99%

“…Combining the CRISPR-Cas12a/Cas13a with microfluidic devices, the detection methods are compatible with the characteristics of rapidity, high sensitivity, low cost, automation and on-site. While microfluidics also brings about new challenges, such as the possible cross-reactivity by diffusion ( Bruch et al, 2021 ), the requirements of appropriately designed channel volume for scale-up ( Ramachandran et al, 2020 ), off-chip manual steps such as sample lysis, nucleic acid extraction as well as nucleic acid amplification ( Ramachandran et al, 2020 ). To solve some of those challenges, futuristic work could be directed towards the development of fully automated microfluidic assays via integration process ( Chen et al, 2020b ).…”

Section: Challenges and Perspectivesmentioning

confidence: 99%

CRISPR-Cas based virus detection: Recent advances and perspectives

Yin¹,

Man²,

et al. 2021

Biosensors and Bioelectronics

144

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Viral infections are one of the most intimidating threats to human beings. One convincing example is the coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2. Rapid, sensitive, specific and field-deployable identification of causal viruses is critical for disease surveillance, control and treatment. The shortcomings of current methods create an impending need for developing novel biosensing platforms. CRISPR-Cas systems, especially CRISPR-Cas12a and CRISPR-Cas13a, characterized by their sensitivity, specificity, high base resolution and programmability upon nucleic acid recognition, have been repurposed for molecular diagnostics, surging a new path forward in biosensing. They, as the core of some robust diagnostic tools, are revolutionizing the way that virus can be detected. This review focuses on recent advances in virus detection with CRISPR-Cas systems especially CRISPR-Cas12a/Cas13a. We started with a short introduction to CRISPR-Cas systems and the properties of Cas12a and Cas13a effectors, and continued with reviewing the current advances of virus detection utilizing CRISPR-Cas systems. The significance and advantages of such methods were then discussed. Finally, the challenges and perspectives were proposed. We tried to provide readers with a concise profile of emerging and fast-expanding CRISPR-Cas based biosensing technology, and highlighted its potential applications in a range of scenarios with regard to virus detection.

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“…Examining the abundance of several components of interest in the same patient’s sample carries the development of diagnostic methods and devices for the multiplexing approach. However, the number of the available Cas protein types and the cross-reaction among different Cas13a proteins can limit the application of Cas13a in multiplex detection [ 63 ]. Hence, a multichannel microfluidic chip approach could overcome this inconvenience.…”

Section: Electrochemical Mirna Detection Based On Crispr-cas13mentioning

confidence: 99%

“…In this regard, more recently, under the same principle used for the EM-CRISPR-Cas13a system, Bruch et al designed and implemented different multiplexed versions of their electrochemical microfluidic biosensor by dividing the channels into subsections, creating four novel chip designs for the amplification-free and simultaneous quantification of up to eight miRNAs through the novel system named CRISPR-Biosensor X. The above can be achieved from a single clinical sample using only one effector protein without changing the sensor or measurement setup [ 63 ].…”

Section: Electrochemical Mirna Detection Based On Crispr-cas13mentioning

confidence: 99%

CRISPR/Cas13-Based Approaches for Ultrasensitive and Specific Detection of microRNAs

Granados-Riverón

Aquino‐Jarquín

2021

Cells

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MicroRNAs (miRNAs) have a prominent role in virtually every aspect of cell biology. Due to the small size of mature miRNAs, the high degree of similarity between miRNA family members, and the low abundance of miRNAs in body fluids, miRNA expression profiling is technically challenging. Biosensors based on electrochemical detection for nucleic acids are a novel category of inexpensive and very sensitive diagnostic tools. On the other hand, after recognizing the target sequence, specific CRISPR-associated proteins, including orthologues of Cas12, Cas13, and Cas14, exhibit collateral nonspecific catalytic activities that can be employed for specific and ultrasensitive nucleic acid detection from clinically relevant samples. Recently, several platforms have been developed, connecting the benefits of enzyme-assisted signal amplification and enzyme-free amplification biosensing technologies with CRISPR-based approaches for miRNA detection. Together, they provide high sensitivity, precision, and fewer limitations in diagnosis through efficient sensors at a low cost and a simple miniaturized readout. This review provides an overview of several CRISPR-based biosensing platforms that have been developed and successfully applied for ultrasensitive and specific miRNA detection.

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CRISPR-powered electrochemical microfluidic multiplexed biosensor for target amplification-free miRNA diagnostics

Cited by 136 publications

References 18 publications

A chemical-enhanced system for CRISPR-Based nucleic acid detection

A chemical-enhanced system for CRISPR-Based nucleic acid detection

CRISPR-Cas based virus detection: Recent advances and perspectives

CRISPR/Cas13-Based Approaches for Ultrasensitive and Specific Detection of microRNAs

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