Related Experiment Video
Updated: Jul 20, 2026
DNA Virus Detection System Based on RPA-CRISPR/Cas12a-SPM and Deep Learning
Published on: May 10, 2024
Rapid and visual detection of Mycoplasma pneumoniae using a novel ERA-CRISPR/Cas12a-based lateral flow assay
1Changzhou Children's Hospital Affiliated to Nantong University, Changzhou, Jiangsu Province 213003 China; Changzhou Medical Center, Nanjing Medical University, Changzhou, Jiangsu Province 213003 China.
Objective:
Mycoplasma pneumoniae (MP) is a leading cause of community-acquired pneumonia in children. Conventional detection methods often lack the speed or accessibility required for point-of-care testing (POCT). This study aims to develop a rapid, highly sensitive, and instrument-free diagnostic platform integrating Enzymatic Rapid Amplification (ERA), CRISPR/Cas12a cleavage, and Lateral Flow Assay (LFA) for visual MP detection.
Methods:
Specific primers targeting the conserved P1 adhesin gene of MP were screened for ERA efficiency. A CRISPR/Cas12a system was designed to specifically recognize the ERA amplicons, triggering the trans-cleavage of a reporter probe. The results were visually interpreted using LFA strips. The assay's limit of detection (LoD), specificity against six common respiratory pathogens, and clinical performance on 80 throat swab samples were evaluated.
Results:
The optimized ERA-CRISPR/Cas12a-LFA assay can be completed within 40 minutes. The LoD was established at 200 copies/mL. Specificity testing showed no cross-reactivity with S. pneumoniae, H. influenzae, or other tested pathogens. In clinical validation (n = 80), the assay demonstrated a sensitivity of 96.23% and specificity of 100% compared to qPCR, with a Kappa value of 0.945.
Conclusion:
The established ERA-CRISPR/Cas12a-LFA method offers a rapid, sensitive, and specific alternative for MP screening. Its minimal equipment requirements make it highly suitable for resource-limited settings and primary care clinics.
Insights
A new diagnostic platform combines Enzymatic Rapid Amplification (ERA), CRISPR/Cas12a, and Lateral Flow Assay (LFA) for rapid, instrument-free detection of Mycoplasma pneumoniae (MP). This method offers high sensitivity and specificity for MP screening, ideal for point-of-care testing.
Area of Science:
- Molecular Diagnostics
- Biotechnology
- Infectious Disease Detection
Background:
- Mycoplasma pneumoniae (MP) is a primary cause of pediatric community-acquired pneumonia.
- Current MP detection methods often lack the speed and accessibility needed for point-of-care testing (POCT).
Purpose of the Study:
- To develop a rapid, sensitive, and instrument-free diagnostic platform for visual MP detection.
- Integrate Enzymatic Rapid Amplification (ERA), CRISPR/Cas12a, and Lateral Flow Assay (LFA) for a novel MP diagnostic approach.
Main Methods:
- Optimized primers targeting the MP P1 adhesin gene for ERA.
- Developed a CRISPR/Cas12a system for specific recognition of ERA amplicons and reporter probe cleavage.
- Utilized Lateral Flow Assay (LFA) for visual result interpretation.
Main Results:
- The ERA-CRISPR/Cas12a-LFA assay provides results within 40 minutes.
- Achieved a limit of detection (LoD) of 200 copies/mL with high specificity against common respiratory pathogens.
- Clinical validation (n=80) showed 96.23% sensitivity and 100% specificity compared to qPCR (Kappa=0.945).
Conclusions:
- The ERA-CRISPR/Cas12a-LFA method is a rapid, sensitive, and specific tool for MP screening.
- Its minimal equipment requirements make it suitable for resource-limited settings and primary care.
- This platform enhances MP diagnostic capabilities at the point of care.
More Related Videos
07:59Rapid and Specific Detection of Acinetobacter baumannii Infections Using a Recombinase Polymerase Amplification/Cas12a-based System
Published on: April 25, 2025
07:46On-site DNA Detection of Trypanosomatid Parasites and Nosema ceranae Through Alkaline Lysis Coupled to RPA/CRISPR/Cas12a System
Published on: July 18, 2025
