Microdroplet Digital PCR for Wide-Range Multiplex Quantification
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Solution Overview
Problem
Conventional digital PCR methods suffer from limited dynamic range, low efficiency, and high workload due to a small number of effective reaction chambers, requiring repetitive detections for multiple target sequences, which increases time and sample consumption.
Innovation Solution
A digital PCR detection apparatus integrating a microdroplet generating device, temperature controlling device, fluorescence signal detecting device, and quantitative analysis device, enabling automatic operations and real-time monitoring of microdroplets for accurate nucleic acid quantification, and a nucleic acid detection microsphere for high-throughput detection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If conventional digital PCR methods use a small number of reaction chambers, then device complexity is reduced, but dynamic range and productivity are limited
Solution Approach 1:
The system segments the reaction system into thousands of independent microdroplets (e.g., 10,000-100,000 droplets per reaction), each serving as an isolated reaction chamber. This segmentation enables the system to achieve a wide dynamic range (5 orders of magnitude) while maintaining manageable device complexity through automated handling of the microdroplet array.
Solution Approach 2:
The patent transitions from conventional single-well or limited-chamber formats to a two-dimensional array of microdroplets on a chip. This dimensional change allows parallel processing of thousands of reactions simultaneously, expanding the dynamic range without proportionally increasing device complexity through integrated automation.
2Adaptability or versatility
If multiple target sequences are detected using conventional digital PCR, then detection comprehensiveness is improved, but workload and time consumption increase due to repetitive detections
Solution Approach 1:
The system employs universal primers that can bind to multiple target sequences simultaneously, allowing a single microdroplet array to detect multiple pathogens or genetic variants in one experiment. This multi-functionality eliminates the need for separate detections for each target, improving both comprehensiveness and efficiency.
Solution Approach 2:
Multiple detection targets are merged into a single reaction system using the same microdroplet array and detection protocol. Different target sequences are detected concurrently within the same experimental run, combining multiple detection functions into one unified process that reduces workload and time consumption.
3Measurement precision
If conventional digital PCR performs successive detections for multiple targets, then detection accuracy is maintained, but time and sample consumption increase
Solution Approach 1:
The system performs continuous parallel detection of multiple target sequences within a single experimental run. Instead of successive discrete detections, all targets are detected simultaneously in continuous parallel operations across the microdroplet array, maintaining accuracy while eliminating idle time between detections.
Solution Approach 2:
The microdroplet array is prepared in advance with all necessary reagents and conditions for detecting multiple targets. This preliminary preparation allows all detections to proceed simultaneously without sequential setup time, reducing total detection time while preserving the accuracy of individual measurements.
4Productivity
If manual operations are used in digital PCR, then device complexity is reduced, but working efficiency decreases
Solution Approach 1:
The system incorporates automated liquid handling, temperature control, and data analysis functions that perform operations without manual intervention. The microdroplet array system self-manages sample distribution, reaction conditions, and result interpretation, improving working efficiency while the integrated design keeps device complexity manageable through consolidation of functions.
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
Enhances working efficiency, accuracy, and sensitivity by allowing simultaneous detection of multiple target sequences with reduced workload and cost, while expanding the dynamic detection range to 5 orders of magnitude using fewer microdroplets.
Implementation Method 1
microdroplet generating device configured to microdropletize a nucleic acid amplification reaction liquid into a plurality of microdroplets
Implementation Method 2
the sample is dispersed to form a plurality of reaction units in the form of water-in-oil
Implementation Method 3
the plurality of microdroplets can be transferred to the temperature controlling device to undergo a temperature cycling to achieve a nucleic acid amplification
Implementation Method 4
undergo a temperature cycling to achieve a nucleic acid amplification
Implementation Method 5
The fluorescence signal detecting device is disposed opposite to the temperature controlling device and configured to photographically detect the plurality of microdroplets after the nucleic acid amplification
Data Source
AI summary
The present application provides a digital PCR detection apparatus, a digital PCR quantitative detection method, a multi-volume digital PCR quantitative analysis method, a digital PCR detection method, a nucleic acid detection microsphere, a preparation method of the nucleic acid detection microsphere, a kit, and a high-throughput nucleic acid detection method.