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Both aerosol and primer dimer breakdown for straightforward genotyping based on an integrated immunochromatographic
Xiaonan Liu1, Jiaxing Zhang2, Kai Hua2
1School of Forensic Medicine, Shanxi Medical University, Taiyuan, 030001, China; College of Life Sciences, Northwest University, Xi'an, 710069, China; Shanxi Key Laboratory of Forensic Medicine, Jinzhong, 030600, China.
Talanta
|December 1, 2024
Summary
This study presents a novel immunochromatographic biosensor for rapid and accurate genotyping. The developed method effectively overcomes false positives from aerosol and primer dimers, enabling point-of-care diagnostics.
Area of Science:
- Biomedical Engineering
- Molecular Diagnostics
- Point-of-Care Testing
Background:
- Genotyping is crucial for disease prevention and treatment, but current point-of-care methods using loop-mediated isothermal amplification (LAMP) and lateral flow often suffer from false positives due to aerosol contamination and primer dimers.
- These limitations hinder the practical application of rapid genotyping technologies in clinical settings.
Purpose of the Study:
- To develop an integrated immunochromatographic biosensor that overcomes the limitations of aerosol contamination and primer dimers in LAMP-based genotyping.
- To enable rapid, accurate, and visually interpretable genotyping at the point-of-care.
Main Methods:
- An integrated immunochromatographic biosensor was developed, incorporating digoxigenin-labeled dUTP into LAMP to allow naked-eye analysis and aerosol elimination via uracil DNA glycosylase.
- Primer dimer interference was resolved by utilizing bio-labeled deoxyribonucleotide and oligonucleotide that do not couple for signal generation.
- Gold magnetic nanoparticles were synthesized as nanoprobe for quantifiable magnetic signal detection, eliminating subjective bias.
Main Results:
- The biosensor successfully eliminated both aerosol contamination and primer dimer interference, addressing key limitations in current lateral flow-based genotyping strategies.
- Genotyping of the C677T polymorphism in methylenetetrahydrofolate reductase was achieved with high accuracy using non-invasive saliva samples and minimal genomic DNA (1 ng).
- Accurate results were obtained within 25 minutes using a simple heater, demonstrating the system's speed and simplicity.
Conclusions:
- The developed immunochromatographic biosensor offers a robust solution for accurate and reliable genotyping at the point-of-care.
- This technology effectively mitigates common sources of false positives, enhancing the practical utility of rapid diagnostic tests.
- The biosensor holds significant potential for facilitating precision medicine through accessible and efficient genetic analysis.
