Reciprocal-Flow PCR Microchannel for Low-Interference Multiplex Detection
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Conventional real-time PCR methods face challenges in multiplex PCR due to fluorescence wavelength interference and require slowing down liquid delivery to avoid interference, leading to inefficiencies in speed and accuracy.
Innovation Solution
A reciprocal-flow-type nucleic acid amplification method that performs thermal cycling by reciprocating a sample liquid between two spatially separated temperature zones in a microchannel, using a microblower for liquid delivery and measuring fluorescence intensity at predetermined points in each zone to minimize noise and enhance speed.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional real-time PCR methods are used for multiplex PCR, then fluorescence detection can be performed, but fluorescence wavelength interference occurs and liquid delivery must be slowed down to avoid interference
Solution Approach 1:
The patent divides the PCR amplification process into two spatially separated temperature zones: a denaturation temperature zone and an elongation-annealing temperature zone. By segmenting the reaction process spatially and using separate fluorescence detection for each zone, the method eliminates fluorescence wavelength interference that occurs in conventional multiplex PCR, while maintaining high amplification speed without requiring slowed liquid delivery.
Solution Approach 2:
The patent introduces an intermediate channel that connects the denaturation temperature zone and the elongation-annealing temperature zone. This intermediate channel serves as a mediator that allows fluorescently labeled primers to be introduced at the denaturation zone while the actual amplification occurs in the elongation-annealing zone, separating the detection function from the amplification function and eliminating interference between multiple fluorescent wavelengths.
2Measurement precision
If liquid delivery is slowed down to avoid fluorescence interference, then measurement precision improves, but the overall PCR operation time increases
Solution Approach 1:
By segmenting the PCR process into separate temperature zones with dedicated functions, the patent allows liquid delivery to proceed at high speed through the intermediate channel without causing fluorescence interference. The denaturation zone handles primer introduction while the elongation-annealing zone handles amplification and detection, eliminating the need to slow down liquid delivery for accurate measurement.
Solution Approach 2:
The patent enables continuous high-speed liquid delivery through the intermediate channel while maintaining accurate fluorescence intensity measurement. The spatial separation allows the liquid flow to continue without interruption or speed reduction, as the fluorescence detection occurs in the elongation-annealing zone where interferenc is eliminated by the zone architecture itself.
3Productivity
If multiple gene regions are amplified simultaneously in multiplex PCR, then productivity improves, but fluorescence wavelength interference increases measurement complexity
Solution Approach 1:
The patent applies segmentation by creating separate functional zones for different aspects of the PCR process. The denaturation temperature zone handles primer introduction and the elongation-annealing temperature zone handles amplification and fluorescence detection. This spatial segmentation simplifies the detection system for multiplex PCR by eliminating cross-wavelength interference, allowing multiple gene regions to be amplified simultaneously with reduced measurement complexity.
Solution Approach 2:
The intermediate channel acts as an intermediary that separates the primer introduction function from the amplification and detection functions. This allows multiple fluorescently labeled primers to be introduced without their signals interfering with each other during detection, simplifying the fluorescence detection system while maintaining the ability to amplify multiple gene regions simultaneously.
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 fast and accurate real-time PCR with reduced noise, enabling simultaneous amplification of multiple gene regions without interference, thus improving the speed and precision of multiplex PCR.
Implementation Method 1
using a microblower for liquid delivery
Implementation Method 2
measuring fluorescence intensity at predetermined points in each zone
Implementation Method 3
two spatially separated temperature zones in a microchannel
Data Source
AI summary
The invention provides a reciprocal-flow-type nucleic acid amplification method performing thermal cycling by reciprocating a sample liquid between a denaturation temperature zone and an elongation-annealing temperature zone with a connected microchannel including at least a curved channel corresponding to the denaturation temperature zone, a curved channel corresponding to the elongation-annealing temperature zone, a linear or curved intermediate channel that connects the aforementioned curved channels, and a connector to connect to a liquid delivery mechanism for enabling movement of the sample liquid. The method includes moving the sample liquid in the channel by the liquid delivery mechanism that is open to atmospheric pressure when liquid delivery is stopped, and measuring a fluorescence intensity for each thermal cycle at a predetermined point on the channel corresponding to the denaturation temperature zone and at a predetermined point on the channel corresponding to the elongation-annealing temperature zone to perform real-time PCR.