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Engineering guidelines for CRISPR diagnostics

delete2025-11-03
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PRE
AI
A
Alexandre S. Avaro
J
Juan G. Santiago *
DOI：10.1039/D5CC04206Bdelete
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Abstract

Abstract

En 中文
This Feature Article reviews engineering guidelines for the design of CRISPR assays; including experimentally validated theoretical models and recommendations for experimental research practice and reporting. First; the state of the art of CRISPR kinetics studies is reviewed. Then presented is a summary of the existence and persistence of widespread gross errors in reports of kinetic rate constants of CRISPR-Cas enzymes; as well as the fact that many CRISPR studies provide insufficient data to check for consistency or assess calibration. Proper experimental procedures including signal calibration are critical to the assessment; design; and future development of CRISPR kinetics assays and CRISPR diagnostics. This review then presents guidelines for the calibration of fluorescence-based CRISPR assays. Fluorescence is the most common detection modality; and incorrect calibration is implicated in high-profile; gross errors in the field. Also presented is a review of enzymatic kinetic rates and reporter molecule degradation as the major factor limiting CRISPR assay sensitivity. Lastly; progress in; and criticism of; microfluidic applications of CRISPR assays is summarized.

Journal

Chemical Communications cover
Chemical Communications
IF:
4.2
Papers:
5.9W
Citations:
16.5W

Organization

No organization information available