Nanozyme-catalysed CRISPR assay for preamplification-free detection of non-coding RNAs

被引:142
作者
Broto, Marta [1 ,2 ]
Kaminski, Michael M. [3 ,4 ,5 ]
Adrianus, Christopher [1 ,2 ]
Kim, Nayoung [1 ,2 ]
Greensmith, Robert [3 ,4 ]
Dissanayake-Perera, Schan [1 ,2 ]
Schubert, Alexander J. [3 ,4 ,5 ]
Tan, Xiao [6 ,7 ,8 ,9 ,10 ]
Kim, Hyemin [1 ,2 ]
Dighe, Anand S. [8 ,11 ]
Collins, James J. [6 ,9 ,10 ,12 ]
Stevens, Molly M. [1 ,2 ]
机构
[1] Imperial Coll London, Dept Mat, Dept Bioengn, London, England
[2] Imperial Coll London, Inst Biomed Engn, London, England
[3] Helmholtz Assoc, Berlin Inst Med Syst Biol, Max Delbruck Ctr Mol Med, Berlin, Germany
[4] Charite Univ Med Berlin, Dept Nephrol & Med Intens Care, Berlin, Germany
[5] Berlin Inst Hlth, Berlin, Germany
[6] Harvard Univ, Wyss Inst Biol Inspired Engn, Boston, MA 02115 USA
[7] Massachusetts Gen Hosp, Div Gastroenterol, Boston, MA 02114 USA
[8] Harvard Med Sch, Boston, MA 02115 USA
[9] MIT, Inst Med Engn & Sci, 77 Massachusetts Ave, Cambridge, MA 02139 USA
[10] MIT, Dept Biol Engn, 77 Massachusetts Ave, Cambridge, MA 02139 USA
[11] Massachusetts Gen Hosp, Dept Pathol, Boston, MA 02114 USA
[12] Broad Inst MIT & Harvard, Infect Dis & Microbiome Program, Cambridge, MA 02142 USA
基金
英国工程与自然科学研究理事会;
关键词
NUCLEIC-ACID DETECTION; AMPLIFICATION;
D O I
10.1038/s41565-022-01179-0
中图分类号
TB3 [工程材料学];
学科分类号
0805 ; 080502 ;
摘要
The combination of catalytic platinum particles, nanozymes and a CRISPR-based reaction allows for the quantification of non-coding RNAs at the picomolar range. This assay, CrisprZyme, has a colorimetric readout and works at room temperature without preamplification. CRISPR-based diagnostics enable specific sensing of DNA and RNA biomarkers associated with human diseases. This is achieved through the binding of guide RNAs to a complementary sequence that activates Cas enzymes to cleave reporter molecules. Currently, most CRISPR-based diagnostics rely on target preamplification to reach sufficient sensitivity for clinical applications. This limits quantification capability and adds complexity to the reaction chemistry. Here we show the combination of a CRISPR-Cas-based reaction with a nanozyme-linked immunosorbent assay, which allows for the quantitative and colorimetric readout of Cas13-mediated RNA detection through catalytic metallic nanoparticles at room temperature (CrisprZyme). We demonstrate that CrisprZyme is easily adaptable to a lateral-flow-based readout and different Cas enzymes and enables the sensing of non-coding RNAs including microRNAs, long non-coding RNAs and circular RNAs. We utilize this platform to identify patients with acute myocardial infarction and to monitor cellular differentiation in vitro and in tissue biopsies from prostate cancer patients. We anticipate that CrisprZyme will serve as a universally applicable signal catalyst for CRISPR-based diagnostics, which will expand the spectrum of targets for preamplification-free, quantitative detection.
引用
收藏
页码:1120 / +
页数:9
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