Functional DNA Regulated CRISPR-Cas12a Sensors for Point-of-Care Diagnostics of Non-Nucleic-Acid Targets

Xiong, Ying; Zhang, Jingjing; Yang, Zhenglin; Mou, Quanbing; Ma, Yuan; Xiong, Yonghua; Lu, Yi

doi:10.1021/jacs.9b09211

Cited by 474 publications

(343 citation statements)

References 59 publications

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“…This method has been used for fluorescent, colorimetric and electronic read-out systems and could detect nucleic acids at as low as femtomolar concentrations in a diversity of sample matrixes. Besides DNA and RNA sensing, detection of proteins and small (inorganic) molecules was also enabled by CRISPR-based sensing ( Dai et al, 2019 ; Liang et al, 2019 ; Xiong et al, 2020 ).…”

Section: Conclusion and Future Outlookmentioning

confidence: 99%

Point-of-care CRISPR/Cas nucleic acid detection: Recent advances, challenges and opportunities

Dongen

Berendsen

Steenbergen

et al. 2020

Biosensors and Bioelectronics

257

191

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With the trend of moving molecular tests from clinical laboratories to on-site testing, there is a need for nucleic acid based diagnostic tools combining the sensitivity, specificity and flexibility of established diagnostics with the ease, cost effectiveness and speed of isothermal amplification and detection methods. A promising new nucleic acid detection method is Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-associated nuclease (Cas)-based sensing. In this method Cas effector proteins are used as highly specific sequence recognition elements that can be combined with many different read-out methods for on-site point-of-care testing. This review covers the technical aspects of integrating CRISPR/Cas technology in miniaturized sensors for analysis on-site. We start with a short introduction to CRISPR/Cas systems and the different effector proteins and continue with reviewing the recent developments of integrating CRISPR sensing in miniaturized sensors for point-of-care applications. Finally, we discuss the challenges of point-of-care CRISPR sensing and describe future research perspectives.

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Section: Conclusion and Future Outlookmentioning

confidence: 99%

Point-of-care CRISPR/Cas nucleic acid detection: Recent advances, challenges and opportunities

Dongen

Berendsen

Steenbergen

et al. 2020

Biosensors and Bioelectronics

257

191

View full text Add to dashboard Cite

show abstract

“…DNA sequences of the HCR system were designed according to the literature and modified to include an aptamer region 39 . In preparation, H1 and H2 were heated separately to 95 °C for 2 min and then cooled to room temperature for 1 h. Then, H0, H1 and H2 were mixed together at concentrations of 0.5, 2 and 2 μM, respectively, in reaction buffer (100 mM Na 2 HPO 4 , 1 mM NaCl and 5 mM EDTA, pH 7.0) and incubated at 37 °C for 1 h. Agarose gels (2%) and PAGE gels were run to confirm the formation of HCR products.…”

Section: Methodsmentioning

confidence: 99%

An ultrasensitive hybridization chain reaction-amplified CRISPR-Cas12a aptasensor for extracellular vesicle surface protein quantification

Xing¹,

Lù²,

Huang³

et al. 2020

Theranostics

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Tumor-derived extracellular vesicle (TEV) protein biomarkers facilitate cancer diagnosis and prognostic evaluations. However, the lack of reliable and convenient quantitative methods for evaluating TEV proteins prevents their clinical application. Methods: Here, based on dual amplification of hybridization chain reaction (HCR) and CRISPR-Cas12a, we developed the apta-HCR-CRISPR assay for direct high-sensitivity detection of TEV proteins. The TEV protein-targeted aptamer was amplified by HCR to produce a long-repeated sequence comprising multiple CRISPR RNA (crRNA) targetable barcodes, and the signals were further amplified by CRISPR-Cas12a collateral cleavage activities, resulting in a fluorescence signal. Results: The established strategy was verified by detecting the TEV protein markers nucleolin and programmed death ligand 1 (PD-L1). Both achieved limit of detection (LOD) values as low as 10 2 particles/µL, which is at least 10 4 -fold more sensitive than aptamer-ELISA and 10 2 -fold more sensitive than apta-HCR-ELISA. We directly applied our assay to a clinical analysis of circulating TEVs from 50 µL of serum, revealing potential applications of nucleolin + TEVs for nasopharyngeal carcinoma cancer (NPC) diagnosis and PD-L1 + TEVs for therapeutic monitoring. Conclusion: The platform was simple and easy to operate, and this approach should be useful for the highly sensitive and versatile quantification of TEV proteins in clinical samples.

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“…This strategy allows the enrollment of more functions such as signal amplification and information processing in the bioanalysis, therefore is greatly adopted in different configurations. For example, the Lu group [303] reported a method of using aptamers and DNAzymes as recognition modules and the CRISPR-Cas systems for signal transduction modules with signal amplification ability. Combination of these two modules enables the quick and sensitive detection on a broad range of targets.…”

Section: Signal Recognition and Transduction Strategiesmentioning

confidence: 99%

Nucleic Acids Analysis

Zhao

Zuo

et al. 2020

Sci. China Chem.

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In recent years, multi-modal entity linking (MEL) has garnered increasing attention in the research community due to its significance in numerous multi-modal applications. Video, as a popular means of information transmission, has become prevalent in people's daily lives. However, most existing MEL methods primarily focus on linking textual and visual mentions or offline videos's mentions to entities in multi-modal knowledge bases, with limited efforts devoted to linking mentions within online video content. In this paper, we propose a task called Online Video Entity Linking (OVEL), aiming to establish connections between mentions in online videos and a knowledge base with high accuracy and timeliness. To facilitate the research works of OVEL, we specifically concentrate on live delivery scenarios and construct a live delivery entity linking dataset called LIVE. Besides, we propose an evaluation metric that considers timelessness, robustness, and accuracy. Furthermore, to effectively handle OVEL task, we leverage a memory block managed by a Large Language Model and retrieve entity candidates from the knowledge base to augment LLM performance on memory management. The experimental results prove the effectiveness and efficiency of our method.

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Functional DNA Regulated CRISPR-Cas12a Sensors for Point-of-Care Diagnostics of Non-Nucleic-Acid Targets

Cited by 474 publications

References 59 publications

Point-of-care CRISPR/Cas nucleic acid detection: Recent advances, challenges and opportunities

Point-of-care CRISPR/Cas nucleic acid detection: Recent advances, challenges and opportunities

An ultrasensitive hybridization chain reaction-amplified CRISPR-Cas12a aptasensor for extracellular vesicle surface protein quantification

Nucleic Acids Analysis

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