One-Pot Pathogen Detection System Resolving Speed-Reliability Trade-off
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Solution Overview
Problem
Current pathogen detection methods, such as loop-mediated isothermal amplification (LAMP), face challenges with false positives and require lengthy reaction times, especially in resource-limited settings, and existing real-time detection methods are prone to errors.
Innovation Solution
A one-pot detection system combining colorimetric, fluorimetric, and lateral flow assays (LFA) to provide multiple readouts, using a reaction mixture with primers, enzymes, and indicators for real-time nucleic acid amplification, allowing for concurrent immunochromatographic assays to validate results quickly.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Speed
If LAMP detection method is used with colorimetric or fluorometric indicators for real-time detection, then detection speed is improved, but false positive rate increases
Solution Approach 1:
The patent combines LAMP amplification with LFA detection in a one-pot system. The LAMP reaction generates amplicons that are simultaneously detected by colorimetric/fluorometric indicators and captured by antibodies on the LFA strip. This merging allows real-time monitoring while maintaining high specificity through the immunochromatographic validation step, resolving the contradiction between speed and reliability.
2Reliability
If LFA is used to detect LAMP result for high specificity, then false positive rate is reduced, but reaction time increases due to pre-defined maximum incubation duration
Solution Approach 1:
The patent implements continuous monitoring of the LAMP reaction through colorimetric or fluorometric indicators throughout the incubation period. This allows the system to detect amplification events as they occur, enabling the LFA validation step to be performed at the optimal moment rather than waiting for a pre-defined maximum incubation time, thus maintaining high specificity while reducing overall reaction time.
3Measurement precision
If multiple detection methods are combined in a single reaction, then diagnostic accuracy is improved, but system complexity increases
Solution Approach 1:
The patent designs a universal one-pot system where a single reaction mixture serves multiple functions: LAMP amplification, colorimetric detection, fluorometric detection, and LFA substrate generation. By making the reaction system multi-functional, the patent achieves high diagnostic accuracy through multiple readouts without proportionally increasing system complexity, as all detection methods share the same amplification platform.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This integrated system reduces false positives and reaction time, offering a highly specific and efficient diagnostic tool suitable for both laboratory and point-of-care settings, with the ability to detect pathogens like MPXV and SARS-CoV-2 with high accuracy and speed.
Implementation Method 1
a colorimetric indicator responsive to a change brought by a potential positive nucleic acid amplification in the reaction mixture
Implementation Method 2
a fluorescent indicator responsive to a presence of amplicons from an amplification reaction of each of target sequences
Implementation Method 3
The antigen-antibody reaction will capture the target amplicons containing the hapten, which are highly specific
Data Source
AI summary
The present disclosure provides a LAMP-based, one-pot pathogen detection system and method integrating a high specificity assay for identifying true positive samples in an analyte before a whole course of LAMP reaction completes, in order to shorten the reaction time and reduce the likelihood of false positive results due to non-specific amplification or signal error (e.g., sample has too high pH) in the LAMP reaction. The present invention integrates an immunochromatographic assay such as lateral flow assay (LFA) in result interpretation from colorimetric and/or fluorimetric aspects of the LAMP reaction in order to enable a real-time pathogen detection, thereby improving the efficiency thereof with high specificity. The present detection system employs a single reaction system to enable multiple result interpretation, and the LAMP reaction can be carried out in a simple platform capable of generating constant heat.