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Cell-based fluorescent microsphere incorporated with carbon dots as a sensitive immunosensor for the rapid detection
Yong Zhao1, Yanzhao Li2, Pingping Zhang1
1State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing, 100071, China; Beijing Key Laboratory of POCT for Bioemergency and Clinic, Beijing, 100071, China.
Abstract:
The rapid and early detection of foodborne pathogens in contaminated food is important for ensuring food safety and quality. In this study, a highly sensitive fluorescent immunosensor was developed to detect Escherichia coli O157:H7 in milk, by using microspheres labeled with carbon dots (CDs). The CDs-microspheres were prepared with Staphylococcus aureus cells as the carrier to incorporate CDs particles. Characterization of the microsphere revealed strong intensity, good stability and high uniformity in fluorescence. With Staphylococcal Protein A (SPA) on the surface of S. aureus cells, the microsphere could be easily coupled with various antibodies (e.g., immunoglobulin G). In combination with the immunomagnetic beads technique, a CDs-microsphere immunosensor was established for the specific detection of E. coli O157:H7 in milk. The limit of detection for E. coli O157:H7 is 2.4 × 102 colony-forming unit (CFU)/mL, comparable to that of real-time PCR methods. Milk samples spiked with E. coli O157:H7 at concentrations from 2.4 × 102 to 2.4 × 107 CFU/mL could be detected within 30 min. The coefficients of variation of the intra-assay tests were less than 10%, indicating a good repeatability. Moreover, the method was able to detect trace amounts of E. coli O157:H7 (<10 CFU) in real milk samples, with a 100% (10/10) accuracy after bacterial enrichment. This CDs-microsphere immunosensor shows considerable potential as a rapid and sensitive tool to detect pathogens in milk and other foods.
Insights
A new fluorescent immunosensor using carbon dots (CDs) microspheres can rapidly detect Escherichia coli O157:H7 in milk. This sensitive method offers accurate pathogen detection for improved food safety.
Area of Science:
- Food safety and quality assurance
- Biosensor development
- Microbiology
Background:
- Rapid and early detection of foodborne pathogens is crucial for public health.
- Escherichia coli O157:H7 is a significant foodborne pathogen causing severe illness.
- Existing detection methods may lack sensitivity or speed for routine screening.
Purpose of the Study:
- To develop a highly sensitive fluorescent immunosensor for detecting Escherichia coli O157:H7 in milk.
- To utilize carbon dots (CDs) labeled microspheres for enhanced detection capabilities.
- To establish a rapid and reliable method for pathogen screening in food matrices.
Main Methods:
- Preparation of carbon dots (CDs)-microspheres using Staphylococcus aureus as a carrier.
- Characterization of microsphere properties including fluorescence intensity, stability, and uniformity.
- Coupling of antibodies to the microspheres via Staphylococcal Protein A (SPA).
- Integration with immunomagnetic beads technique for specific E. coli O157:H7 detection.
Main Results:
- The developed immunosensor demonstrated high fluorescence intensity, stability, and uniformity.
- A limit of detection of 2.4 × 10^2 CFU/mL for E. coli O157:H7 was achieved, comparable to real-time PCR.
- Detection of spiked milk samples was accomplished within 30 minutes.
- Excellent repeatability with intra-assay coefficients of variation <10%.
- Accurate detection of trace amounts (<10 CFU) in real milk samples after enrichment (100% accuracy).
Conclusions:
- The carbon dots-microsphere immunosensor is a sensitive and rapid tool for detecting E. coli O157:H7 in milk.
- This technology shows significant potential for application in broader food safety monitoring.
- The method offers a promising alternative for pathogen detection in various food products.
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