Microfluidic Device With Cleavable Solid Supports for Multiplexed PCR
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Solution Overview
Problem
Current PCR technologies face challenges in simultaneous and quantitative analysis of multiple analytes due to limitations in detection methods, primer and probe set design, and instrumentation, which hinders rapid reconfiguration for emerging diseases and increases complexity in sample preparation and device production.
Innovation Solution
A microfluidic device with a plurality of reaction wells and solid supports where reagents, such as primers for PCR, are attached via labile bonds that can be cleaved using thermal, chemical, or light-based operations, allowing for controlled release of reagents into reaction wells for multiplexed PCR reactions.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If multiplexed PCR reactions are performed using a small number of reaction wells with multiple color probes, then simultaneous analysis of multiple analytes is enabled, but the design and optimization of primer and probe sets becomes extremely challenging and complex
Solution Approach 1:
The patent segments the reagents (primers and probes) by attaching them to individual solid supports (beads) instead of using them in bulk solution. Each bead carries specific reagents, allowing independent control and simplifying the design process. This segmentation transforms the complex multiplexed reaction design into a more manageable bead-based system where reagents are pre-assembled and stored separately.
Solution Approach 2:
The patent introduces solid supports (beads) as intermediary carriers between the reagents and the reaction environment. These beads serve as mediators that hold specific primer and probe sets, enabling controlled release into reaction wells. This intermediary approach simplifies the system by providing a stable platform for reagent delivery and reducing the complexity of designing compatible multi-color probe sets.
2Productivity
If preloading of primer/probe sequences is done using printing-based technology, then high-throughput analysis is enabled, but the time and expense required for device production greatly increases
Solution Approach 1:
Instead of using expensive printing-based technology to load reagents, the patent uses solid supports (beads) as reusable copies or templates that can be manufactured more simply. The beads serve as replicable units that can be produced through standard microsphere fabrication methods, reducing manufacturing complexity and cost while maintaining high-throughput capability.
Solution Approach 2:
The patent changes the physical state and delivery mechanism of reagents from liquid-based printing to solid-based bead attachment. This parameter change enables simpler manufacturing processes while maintaining the ability to deliver reagents in high-throughput fashion. The solid support approach allows for easier scaling and reduced production time compared to traditional printing methods.
3Stability of the object's composition
If reagents are attached to solid supports via stable bonds, then reagent stability is improved, but the ability to control release timing and conditions is reduced
Solution Approach 1:
The patent uses bonds with different strength parameters that can be selectively broken under specific conditions. By choosing bonds with appropriate bond energies, the system achieves stable attachment during storage and transport, but allows controlled release when specific thermal, chemical, or physical conditions are applied during the PCR reaction. This parameter-based control enables both stability and adaptability.
Solution Approach 2:
The patent introduces dynamic control over reagent release by using bonds that can transition from stable to labile states. The bonds are designed to remain stable under normal storage conditions but become labile when specific triggers are applied (heat, chemicals, light). This dynamic property allows the system to adapt between different states: stable for storage, reactive for execution, thereby resolving the contradiction between stability and controllability.
4Productivity
If conventional multiplexed PCR is used, then analysis of 3-12 targets is possible, but the approach becomes cumbersome for significantly larger numbers of targets
Solution Approach 1:
The patent segments reagents onto individual beads, creating a modular system where each bead can be independently designed and attached. This segmentation allows for easy reconfiguration when analyzing new sequences, as beads can be simply replaced or regenerated without redesigning the entire assay. This modular approach scales efficiently from 3-12 to hundreds of targets.
Solution Approach 2:
The patent performs preliminary attachment of reagents to beads during manufacturing, creating pre-assembled reaction units. This preliminary action eliminates the need for complex real-time reconfiguration when analyzing new targets, as the bead-based reagents can be quickly regenerated or exchanged. This approach maintains ease of operation even when scaling to large numbers of targets.
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 enables efficient and controlled release of reagents, reducing primer dimer formation and non-specific interactions, and allows for high-throughput, multiplexed PCR analysis with reduced reagent consumption and simplified device production, facilitating rapid reconfiguration for new assays.
Implementation Method 1
the reagent is configured to be cleaved from the support via a cleaving operation. The cleaving operation may include a thermal operation.
Implementation Method 2
The cleaving operation may include... an application of light to the reagent/support bond.
Data Source
AI summary
A microfluidic device includes a plurality of reaction wells; and a plurality of solid supports, and each of the solid supports has a reagent attached thereto. The reagent is attached to the solid support via a labile reagent/support bond such that the reagent is configured to be cleaved from the support via a cleaving operation.