Immunoassay System Based on the Technology of Time-Resolved Fluorescence Resonance Energy Transfer

Area of Science:

• Biomedical Engineering
• Analytical Chemistry
• Biotechnology

Background:

• Traditional immunoassays face challenges in specificity, sensitivity, and cost-effectiveness.
• Multiplexed detection of biomarkers is crucial for accurate disease diagnosis and monitoring.
• Time-resolved fluorescence resonance energy transfer (TR-FRET) offers a promising approach to overcome these limitations.

Purpose of the Study:

• To develop and validate a novel immunoassay system utilizing TR-FRET technology.
• To enhance assay specificity and sensitivity while reducing detection costs.
• To enable simultaneous detection of multiple biomarkers, including procalcitonin (PCT), C-reactive protein (CRP), and interleukin-6 (IL-6).

Main Methods:

• Designed and constructed an instrument incorporating a xenon flash lamp, photomultiplier tube (PMT), and a filter-switching mechanism for alternate donor and acceptor fluorescence measurement.
• Employed TR-FRET principles, leveraging long-lifetime fluorescence from acceptor probes for enhanced sensitivity.
• Validated the instrument's performance using PCT, CRP, and IL-6 assays across various concentrations.

Main Results:

• The developed instrument demonstrated excellent performance with low noise levels and minimal temperature variations.
• High linearity (R² > 0.999) was achieved for luminescence values and biomarker detection across specified ranges.
• Achieved sensitive quantification limits for PCT (0.096 ng/mL), CRP (2.70 ng/mL), and IL-6 (2.82 ng/mL).

Conclusions:

• The developed TR-FRET immunoassay system is feasible and effective for sensitive, specific, and cost-efficient multiplexed biomarker detection.
• The instrumental and technical solutions provide a robust platform for advanced immunoassays.
• This technology holds potential for improved diagnostic capabilities in clinical settings.