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<p>A novel biosensing platform has been developed that couples the collateral cleavage activity of the CRISPR/Cas13a system with single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS) using gold nanoparticle (AuNP)-DNA reporter probes. In the presence of target microRNA-21, Cas13a is activated and specifically cleaves RNA bases within the DNA-RNA hybrid linkers on AuNPs, preventing their hybridization with biotinylated capture probes on magnetic beads. As a result, the cleaved AuNPs remain in the supernatant and are directly quantified by sp-ICP-MS. The number of detected AuNPs correlates linearly with the concentration of miRNA-21 (y = 0.2537logC<sub>miRNA</sub> + 0.2477 with a correlation coefficient of R<sup>2</sup> = 0.9978), enabling a detection limit as low as 68 fM with excellent single-base mismatch discrimination. The assay demonstrated high recoveries (97–108%) and reproducibility in spiked human serum samples, confirming its reliability in complex matrices. This amplification-free strategy combines the high specificity of CRISPR/Cas13a with the single-particle sensitivity of sp-ICP-MS, providing a robust, quantitative, and versatile platform for miRNA detection with promising potential for early cancer diagnostics and precision medicine.</p>

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Single particle mediated CRISPR/Cas13a ultrasensitive direct detection of miRNA-21 at femtomolar levels without nucleic acid amplification

  • 期刊论文

  • 发表日期：

    2026年2月26日

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  • Nandi Chen,
  • Xiao Sun,
  • Shaoxia Liang,
  • Chong Cao,
  • Xuandi Lai,
  • Chunyan Song,
  • Zhigang Yan,
  • Chenchen Ge

摘要

A novel biosensing platform has been developed that couples the collateral cleavage activity of the CRISPR/Cas13a system with single-particle inductively coupled plasma mass spectrometry (sp-ICP-MS) using gold nanoparticle (AuNP)-DNA reporter probes. In the presence of target microRNA-21, Cas13a is activated and specifically cleaves RNA bases within the DNA-RNA hybrid linkers on AuNPs, preventing their hybridization with biotinylated capture probes on magnetic beads. As a result, the cleaved AuNPs remain in the supernatant and are directly quantified by sp-ICP-MS. The number of detected AuNPs correlates linearly with the concentration of miRNA-21 (y = 0.2537logCmiRNA + 0.2477 with a correlation coefficient of R2 = 0.9978), enabling a detection limit as low as 68 fM with excellent single-base mismatch discrimination. The assay demonstrated high recoveries (97–108%) and reproducibility in spiked human serum samples, confirming its reliability in complex matrices. This amplification-free strategy combines the high specificity of CRISPR/Cas13a with the single-particle sensitivity of sp-ICP-MS, providing a robust, quantitative, and versatile platform for miRNA detection with promising potential for early cancer diagnostics and precision medicine.