Microcarrier Multiplex DNA Mutation Detection Assay
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Solution Overview
Problem
Current methods for detecting DNA mutations associated with colorectal cancer are invasive, time-consuming, and lack uniformity, requiring multiple assays that are expensive and cumbersome, especially when dealing with limited sample volumes from invasive sources.
Innovation Solution
The use of microcarriers encoded with unique identifiers and probes specific for DNA mutations in the KRAS, BRAF, CTNNB1, and APC genes, allowing for multiplex detection through PCR amplification and hybridization, followed by fluorescence imaging for accurate mutation identification.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If multiple singleplex assays are performed separately in different reaction vessels, then each target can be detected with high accuracy, but the device complexity and time consumption increase significantly
Solution Approach 1:
The patent combines multiple singleplex assays into a single multiplex assay by immobilizing different capture agents on the same solid phase surface, allowing simultaneous detection of multiple targets in one reaction vessel, thereby reducing device complexity and time consumption while maintaining detection accuracy
Solution Approach 2:
The solid phase surface is designed with universal functionality to support multiple capture agents and detection molecules simultaneously, enabling a single assay system to perform multiple detection functions for different DNA targets, mutations, or biomarkers
2Measurement precision
If multiple singleplex assays are performed separately, then each target can be analyzed independently, but the time required and sample volume needed increase dramatically
Solution Approach 1:
Multiple assay steps that were previously performed sequentially in separate vessels are merged into a single parallel reaction on the solid phase surface, allowing independent target analysis to occur simultaneously, thereby dramatically reducing total assay time while maintaining analytical precision
Solution Approach 2:
The multiplex assay enables continuous parallel processing of multiple targets without interruption or sequential waiting, maintaining useful analytical action across all targets simultaneously, which eliminates time losses associated with sequential singleplex assays
3Adaptability or versatility
If multiple singleplex assays are conducted in parallel, then comprehensive coverage of multiple targets is achieved, but the required sample volume becomes prohibitively large
Solution Approach 1:
The patent merges multiple assay reactions into a single shared reaction volume on the solid phase surface, allowing comprehensive coverage of multiple targets using a single small sample aliquot, thereby dramatically reducing the total sample volume required compared to parallel singleplex assays
Solution Approach 2:
The solid phase assay system provides universal target capture capability that can simultaneously analyze multiple DNA targets, mutations, or biomarkers in a single reaction, enabling comprehensive target coverage with minimal sample volume requirements
4Ease of manufacture
If separate assays are performed for each target, then reagent costs per target can be optimized, but the total reagent cost and operational complexity increase
Solution Approach 1:
The patent merges multiple assay procedures into a single standardized multiplex protocol, eliminating the need to perform and manage multiple separate assays, thereby simplifying operational procedures and improving throughput while maintaining reagent cost efficiency through shared resources
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 rapid, cost-effective, and accurate multiplex detection of multiple DNA mutations in a single assay, reducing the need for extensive sample volumes and improving diagnostic efficiency for colorectal cancer screening and monitoring.
Implementation Method 1
hybridizing the amplified DNA with at least four probes, said at least four probes comprising one or more probes specific for a DNA mutation
Implementation Method 2
followed by fluorescence imaging for accurate mutation identification
Data Source
AI summary
Provided herein are methods and kits for detecting the presence of DNA mutations in the KRAS, BRAF, CTNNB1, and APC genes. The methods and kits employ microcarriers, each with a probe specific for a DNA mutation in the KRAS, BRAF, CTNNB1, or APC gene and an identifier unique to the probe sequence. Upon isolation and amplification of DNA from a sample, hybridization of amplified DNA with a probe, specific for a DNA mutation, that is coupled to a microcarrier indicates the presence of the DNA mutation in the sample. Since each microcarrier can be identified through detection of the identifier, multiplex screening assays for multiple mutations in each of the KRAS, BRAF, CTNNB1, and APC genes are provided.