dcm-Targeted LAMP Primer Kit for Rapid Gonorrhea Detection
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Current diagnostic methods for Neisseria gonorrhoeae are labor-intensive, time-consuming, require specialized equipment, and are costly, with limited sensitivity and specificity, especially for point-of-care testing (POCT), and existing LAMP methods lack sensitivity and speed for early detection.
Innovation Solution
A set of primers targeting the Neisseria gonorrhoeae DNA cytosine methyltransferase (dcm) gene, combined with the LAMP method, using a specific temperature profile and fluorescent dye for rapid and sensitive detection, allowing for quantitative measurement and low detection limits.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If Real-Time PCR-based assays are used for detecting Neisseria gonorrhoeae, then sensitivity and specificity are improved, but cost, equipment requirements, and analysis time increase significantly
Solution Approach 1:
The patent replaces the complex thermal cycling mechanical system of Real-Time PCR with a simple isothermal amplification system using LAMP (Loop-mediated Isothermal Amplification). This substitution eliminates the need for expensive thermal cyclers while maintaining high detection sensitivity through a different amplification mechanism that operates at constant temperature (65°C), thereby resolving the contradiction between measurement precision and device complexity.
Solution Approach 2:
The patent changes the fundamental operating parameter from cyclic temperature variation (PCR) to constant isothermal condition (LAMP). By maintaining a single temperature of 65°C throughout the amplification process, the method eliminates complex thermal cycling equipment while achieving comparable or superior sensitivity through continuous amplification, thus resolving the contradiction between detection sensitivity and equipment complexity.
2Measurement precision
If Real-Time PCR-based assays are used for detecting Neisseria gonorrhoeae, then sensitivity and specificity are improved, but analysis time and energy consumption increase
Solution Approach 1:
The patent employs periodic addition of reagents (beta-glycerophosphate and MgSO4) at specific time intervals during the isothermal amplification process to optimize reaction efficiency and reduce time-to-positive result. This periodic intervention allows the reaction to progress through distinct phases (exponential amplification followed by linear amplification), achieving rapid detection in 15-30 minutes while maintaining high sensitivity, thereby resolving the contradiction between measurement precision and loss of time.
3Measurement precision
If molecular probes labelled with fluorescent dyes are used for quantitative measurement, then measurement precision is improved, but cost increases significantly
Solution Approach 1:
The patent employs inexpensive fluorescent dyes (such as SYBR Green or EvaGreen) that bind non-specifically to amplified DNA products rather than using expensive sequence-specific molecular probes. These dyes provide sufficient quantitative measurement capability through simple fluorescence intensity correlation with DNA amount, while costing a fraction of probe-based methods. The dyes are added in excess and require no complex optimization, thereby resolving the contradiction between measurement precision and quantity of substance (cost).
4Loss of time
If isothermal LAMP method is used for rapid detection, then analysis time is reduced, but sensitivity and detection limit are insufficient for early detection
Solution Approach 1:
The patent performs preliminary optimization of primer concentrations, Mg2+ concentration, and betaine addition timing to maximize amplification efficiency during the critical early exponential phase. By pre-establishing optimal reaction conditions and adding enhancing reagents (beta-glycerophosphate and MgSO4) at precise intervals, the method achieves ultra-sensitive detection (≤10 copies/reaction) within 15-30 minutes, resolving the contradiction between analysis time and measurement precision through advance preparation and timed intervention.
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
The method achieves rapid detection of Neisseria gonorrhoeae with a detection limit of 10 copies/reaction in under 20 minutes, suitable for POCT, with improved sensitivity and specificity, enabling early targeted therapy.
Implementation Method 1
Isothermal methods, including the LAMP (Loop-mediated isothermal amplification) method, are methods that allow to accelerate the diagnostic process and reduce the cost of energy needed to perform the analysis
Implementation Method 2
fluorescent dye for rapid and sensitive detection, allowing for quantitative measurement
Data Source
AI summary
The invention relates to a set of amplification primers, a method for detecting a sexually transmitted bacterial infection, and a kit for detecting the infection.