Thermal Block Temperature Correction for Nucleic Acid Amplification
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Solution Overview
Problem
Current nucleic acid amplification systems with multiple thermal control blocks face challenges in achieving precise temperature control, particularly for applications like PCR and HRM analysis, where temperature differences between blocks can exceed ±0.5°C, affecting amplification efficiency and reproducibility.
Innovation Solution
A nucleic acid amplification system with individually temperature-controllable thermal blocks, a real-time fluorescence measuring unit, and a temperature correction unit that uses a reference melting temperature to correct temperature absolute values, ensuring uniformity between blocks to ±0.05°C or less by measuring the melting temperature of a calibration sample.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If multiple thermal control blocks are used for parallel nucleic acid amplification, then productivity is improved, but temperature uniformity between blocks deteriorates
Solution Approach 1:
The system performs preliminary temperature calibration by measuring the melting temperature of a calibration sample in each thermal control block before actual amplification. Correction values are calculated and stored in advance, which are then applied during subsequent amplification processes to compensate for temperature deviations and achieve uniform temperature control across multiple blocks.
Solution Approach 2:
The system implements feedback control by measuring the actual melting temperature in each thermal control block, comparing it with the target value, calculating the deviation, and applying correction values to adjust the temperature control. This closed-loop feedback mechanism ensures temperature uniformity is maintained across all blocks during parallel amplification.
2Measurement precision
If conventional temperature measurement probes are used for calibration, then temperature control is achieved, but measurement precision is limited to ±0.25°C
Solution Approach 1:
The system uses a calibration sample with a known melting temperature as an intermediary reference standard. Instead of directly measuring temperature with probes, the system measures the melting temperature of the calibration sample, which serves as a mediator to indirectly determine and correct the actual temperature in each thermal control block, achieving higher precision without complex direct temperature measurement systems.
Solution Approach 2:
The system replaces the mechanical temperature measurement probe calibration method with a fluorescence-based melting temperature measurement approach. By using fluorescence signals to detect the melting temperature of a calibration sample, the system achieves higher measurement precision while avoiding the limitations of conventional probe-based mechanical measurement systems.
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 approach achieves precise temperature uniformity between thermal control blocks, enhancing DNA amplification efficiency and reproducibility, and improving the accuracy of real-time PCR and HRM analysis by minimizing temperature differences.
Implementation Method 1
a plurality of individually temperature-controllable thermal control blocks (1)
Implementation Method 2
a real-time fluorescence measuring unit (3) for performing real-time fluorescence measurement on a sample in a reaction vessel (21)
Implementation Method 3
a melting temperature measurement unit (7) for measuring a melting temperature of a temperature calibration sample (21)
Data Source
AI summary
According to a conventional technique, when a calibrated temperature measuring probe is used for correcting the temperature absolute values of individually temperature-controllable thermal control blocks, a temperature difference of a maximum of 0.5° C. remains between the thermal control blocks. According to the present invention, the melting temperature of a temperature calibration sample housed in a reaction vessel corresponding to each of the temperature control blocks is measured as a measured melting temperature. The measured melting temperature corresponding to each of the thermal control blocks and the reference melting temperature of the temperature calibration sample are compared, and the temperature absolute value of each of the thermal control blocks is corrected based on respective difference values.