Innovative PNA-LB mediated allele-specific LAMP for KRAS mutation profiling on a compact lab-on-a-disc device

Area of Science:

• Biotechnology
• Molecular Diagnostics
• Oncology

Background:

• Personalized medicine requires advanced diagnostic tools for individual patient treatment.
• KRAS mutations are key biomarkers for selecting patients for targeted cancer therapies.
• Accurate and user-friendly detection of KRAS mutations is essential for effective treatment selection.

Purpose of the Study:

• To develop a novel Lab-on-a-Disc system for detecting the KRAS G12D mutation.
• To evaluate the system's precision, accuracy, sensitivity, and specificity.
• To demonstrate the system's superiority over existing mutation detection methods.

Main Methods:

• Utilized a Lab-on-a-Disc platform with peptide nucleic acids-loop backward (PNA-LB) mediated allele-specific loop-mediated isothermal amplification (LAMP).
• Incorporated a robust microfluidic design, precise rotary control, and induction heating for multiplexed LAMP reactions.
• Employed naked-eye observation, RGB image analysis, and time of fluorescence (Tf) for endpoint detection.

Main Results:

• Achieved high preclinical and technical precision, accuracy, and versatility.
• Increased assay sensitivity and specificity by 80% and 90% respectively, by optimizing cutoff values.
• Demonstrated high reproducibility (CV% < 3.5%) and repeatability (CV% < 5.5%) for multiplexed LAMP reactions.
• Preclinical performance was comparable to standard methods using various detection readouts.

Conclusions:

• The novel Lab-on-a-Disc system provides a precise and efficient method for KRAS G12D mutation detection.
• This technology significantly advances the development of tailored treatment strategies for cancer patients.
• The system's features enable accurate, user-friendly, and reproducible mutation analysis for personalized oncology.