Rolling Circle Amplification-Enabled Ultrasensitive Point-of-Care Test Method for Aflatoxin B1 in the Environment and...

Area of Science:

• Analytical Chemistry
• Biotechnology
• Environmental Science

Background:

• Aflatoxin B1 (AFB1) contamination presents a significant health risk, necessitating sensitive detection methods for food safety and environmental monitoring.
• Current immunoassay methods often lack the required sensitivity and are susceptible to complex matrix effects.
• Rolling circle amplification (RCA) offers high specificity and sensitivity for nucleic acid amplification, making it a promising technique.

Purpose of the Study:

• To develop a general, highly sensitive, and reliable point-of-care test (POCT) method for Aflatoxin B1 (AFB1) detection.
• To overcome the sensitivity limitations and matrix effects associated with existing immunoassay techniques for AFB1.
• To establish a versatile platform for AFB1 detection in diverse sample matrices like water, soil, and food.

Main Methods:

• Construction of a general rolling circle amplification-based point-of-care test (RCA-POCT) method.
• Utilized biotinylated antibodies, streptavidin, and biotinylated RCA primers for signal transduction and amplification.
• Employed a fluorescence-based detection system for immunocomplexes in microwells.

Main Results:

• Achieved sensitive detection limits for AFB1: 1.94 fg/mL in irrigation water, 16.3 fg/mL in field soil, and 37.7 fg/mL in peanut samples.
• Demonstrated wide linear ranges for AFB1 detection across different matrices (e.g., 5 × 10⁻⁶ to 5 ng/mL in irrigation water).
• Exhibited satisfactory recovery, specificity, repeatability, and reproducibility, validated against High-Performance Liquid Chromatography (HPLC).

Conclusions:

• The developed RCA-POCT method provides a sensitive and generalizable approach for detecting Aflatoxin B1.
• This method effectively enhances fluorescent signals, addressing sensitivity and matrix effect challenges in AFB1 detection.
• The RCA-assisted detection offers a valuable tool for rapid AFB1 monitoring in environmental and food safety applications.