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<p>Postoperative infections, particularly those caused by methicillin-resistant <i>Staphylococcus aureus</i> (MRSA), pose a severe threat to patients receiving postoperative nursing such as pulmonary thromboendarterectomy for chronic thromboembolic pulmonary hypertension. The need for rapid, accurate, and sensitive MRSA detection in clinical settings remains unmet by conventional culture-based or single-analyte molecular methods, which are often slow, complex, or lack sufficient specificity and sensitivity. Herein, we report a novel, integrated biosensing platform that achieves ultrasensitive and specific MRSA detection by synergistically combining proximity hybridization, hybridization chain reaction (HCR), T7 RNA polymerase-driven transcription, and CRISPR-Cas12a <i>trans</i>-cleavage. The assay initiates with the dual recognition of MRSA surface markers, Protein A and PBP2a, by antibody-conjugated DNA probes. Subsequent proximity hybridization triggers an HCR cascade, forming a long DNA polymer that serves as a template for T7-mediated transcription, generating abundant target-specific crRNAs. These crRNAs guide Cas12a complexes, whose activated <i>trans</i>-cleavage activity produces a strong, quantifiable fluorescent signal. This meticulously orchestrated, multi-stage amplification strategy confers the assay with exceptional performance: a wide dynamic range from 10 to 10⁶ CFU/mL, a low detection limit, and excellent specificity against common interfering pathogens. The method demonstrated high reliability in spiked serum samples, showing strong correlation with the gold-standard culture technique. By seamlessly integrating protein recognition with nucleic acid amplification and CRISPR-based signal generation, this work presents a significant advancement in diagnostic technology for timely and precise monitoring of MRSA infections during postoperative nursing.</p>

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A cascade amplification platform integrating dual-protein recognition and CRISPR-Cas12a for accurate MRSA analysis in postoperative care

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年7月6日

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  • Wenjia Mao,
  • Bin Li,
  • Shangjun Huang

摘要

Postoperative infections, particularly those caused by methicillin-resistant Staphylococcus aureus (MRSA), pose a severe threat to patients receiving postoperative nursing such as pulmonary thromboendarterectomy for chronic thromboembolic pulmonary hypertension. The need for rapid, accurate, and sensitive MRSA detection in clinical settings remains unmet by conventional culture-based or single-analyte molecular methods, which are often slow, complex, or lack sufficient specificity and sensitivity. Herein, we report a novel, integrated biosensing platform that achieves ultrasensitive and specific MRSA detection by synergistically combining proximity hybridization, hybridization chain reaction (HCR), T7 RNA polymerase-driven transcription, and CRISPR-Cas12a trans-cleavage. The assay initiates with the dual recognition of MRSA surface markers, Protein A and PBP2a, by antibody-conjugated DNA probes. Subsequent proximity hybridization triggers an HCR cascade, forming a long DNA polymer that serves as a template for T7-mediated transcription, generating abundant target-specific crRNAs. These crRNAs guide Cas12a complexes, whose activated trans-cleavage activity produces a strong, quantifiable fluorescent signal. This meticulously orchestrated, multi-stage amplification strategy confers the assay with exceptional performance: a wide dynamic range from 10 to 10⁶ CFU/mL, a low detection limit, and excellent specificity against common interfering pathogens. The method demonstrated high reliability in spiked serum samples, showing strong correlation with the gold-standard culture technique. By seamlessly integrating protein recognition with nucleic acid amplification and CRISPR-based signal generation, this work presents a significant advancement in diagnostic technology for timely and precise monitoring of MRSA infections during postoperative nursing.