Multiplex qPCR Panel for Specific Upper Respiratory Pathogen Detection
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Solution Overview
Problem
There is a need for a highly sensitive and specific multiplex assay to detect human metapneumovirus, adenovirus, parainfluenza virus, and rhinovirus/enterovirus in a single test, addressing the challenges of cross-reactivity and complexity in respiratory pathogen detection.
Innovation Solution
A nucleic acid-based method using primer pairs and probes to amplify and detect specific genomic regions of these pathogens in a multiplex qPCR assay, with an internal control for sample integrity, utilizing distinct fluorophores for each pathogen and RNase P as an endogenous marker.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If multiple respiratory pathogens are detected using separate assays, then detection sensitivity and specificity for each pathogen can be maintained, but the complexity of the diagnostic workflow increases and throughput decreases
Solution Approach 1:
The patent combines multiple separate pathogen detection assays into a single multiplex qPCR assay that can simultaneously detect adenovirus, metapneumovirus, parainfluenza virus, and rhinovirus/enterovirus in one reaction well, thereby increasing throughput and simplifying workflow while maintaining detection sensitivity and specificity through pathogen-specific primer and probe designs
Solution Approach 2:
The invention creates a universal multiplex panel that can detect multiple different respiratory pathogens using a single standardized assay platform with common reagents and protocols, allowing the same system to perform multiple detection functions across different pathogen types without requiring separate specialized assays for each pathogen
2Productivity
If a multiplex assay detects multiple pathogens simultaneously, then throughput increases and workflow simplifies, but the risk of cross-reactivity between primers and probes increases
Solution Approach 1:
The patent applies local quality by designing pathogen-specific primer and probe sequences that target unique genomic regions for each pathogen (adenovirus, metapneumovirus, parainfluenza virus, rhinovirus/enterovirus), ensuring that each detection element is highly specific to its intended target and does not cross-react with other pathogens in the multiplex panel
Solution Approach 2:
The invention optimizes specific parameters including primer melting temperatures, probe fluorophore selection, and qPCR cycling conditions to ensure specific amplification and detection of each pathogen while preventing cross-reactivity, with each pathogen detected using distinct fluorophores for clear differentiation
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 provides sensitive and specific detection of multiple respiratory pathogens in a single assay, ensuring high throughput and low cost, with a simple workflow.
Implementation Method 1
The generating of the at least one amplicon includes subjecting the reaction mixture to polymerase chain reaction (PCR) conditions
Implementation Method 2
utilizing distinct fluorophores for each pathogen
Data Source
AI summary
An in vitro method, composition and kit for determining the presence or absence of adenovirus, metapneumovirus, rhinovirus/enterovirus, and parainfluenza in a sample, including providing a reaction mixture containing the sample and at least one primer pair set. The primer pair set includes at least one primer pair A that specifically amplifies a portion of adenovirus genome; at least one primer pair B that specifically amplifies a portion of metapneumovirus genome; at least one primer pair C that specifically amplifies a portion of rhinovirus/enterovirus genome; and at least one primer pair D that specifically amplifies a portion of parainfluenza genome. The reaction mixture is subjected to reaction conditions suitable to amplify targeted nucleic acids, thereby generating at least one amplicon; wherein the presence or absence of at least one amplicon in the sample indicates the presence or absence of adenovirus, metapneumovirus, rhinovirus/enterovirus, and/or parainfluenza in the sample.