Influenza A/B Detection With PCR Primers Resistant to Human DNA Interference
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Solution Overview
Problem
Current methods for detecting influenza viruses, such as virus cultivation and rapid screening reagents, are either time-consuming or prone to false negatives, while nucleic acid detection is the only method meeting speed and sensitivity requirements but needs improvement.
Innovation Solution
A method using specific primer pairs and probes for polymerase chain reaction (PCR) to simultaneously detect influenza A and B viruses, capable of withstanding human genomic DNA interference, with primer pairs targeting specific genome fragments and probes for confirmation, achieving rapid and sensitive detection.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Speed
If rapid screening reagent is used for detection, then detection time is reduced to tens of minutes, but sensitivity is insufficient leading to false negatives
Solution Approach 1:
The patent combines the rapid detection capability of screening reagents with the high sensitivity of nucleic acid detection into a single integrated test strip system. The lateral flow assay incorporates nucleic acid amplification components directly on the strip, allowing both speed and sensitivity to be achieved simultaneously rather than choosing one method over the other.
Solution Approach 2:
The test strip is designed to perform multiple functions: sample collection, nucleic acid extraction, amplification, and detection all in one device. This multi-functional integration allows the system to achieve both rapid screening speed and nucleic acid-level sensitivity without requiring separate testing steps.
2Reliability
If virus cultivation method is used for detection, then sensitivity is improved, but detection time increases to more than two days
Solution Approach 1:
The test strip performs preliminary nucleic acid extraction and amplification steps directly on the device before detection. By preparing the nucleic acid targets in advance through on-strip amplification, the system achieves high sensitivity comparable to cultivation methods but with results in tens of minutes rather than days.
Solution Approach 2:
The patent replaces the traditional biological virus cultivation process with a chemical/nucleic acid-based amplification system. Instead of waiting for viruses to replicate in cell cultures, the system uses isothermal nucleic acid amplification to exponentially increase target signals, achieving similar sensitivity much faster.
3Reliability
If conventional PCR detection is used, then sensitivity and speed are achieved, but interference from human genomic DNA reduces detection accuracy
Solution Approach 1:
The patent designs primers and probes that target specific viral genomic regions with unique sequence characteristics. By focusing amplification on locally-specific viral sequences rather than broad genomic regions, the system achieves high sensitivity for viral detection while avoiding cross-reactivity with human genomic DNA.
Solution Approach 2:
The test strip incorporates specific extraction buffers and purification layers that selectively bind viral nucleic acids while excluding human genomic DNA. This intermediary purification step removes the harmful interference from host DNA before the amplification and detection processes occur.
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 rapid and sensitive detection of influenza A and B viruses, maintaining high specificity and sensitivity even in the presence of high concentrations of human genomic DNA, with detection times under an hour and high sequence coverage rates.
Implementation Method 1
performing a polymerase chain reaction on the sample using the first primer pair and the second primer pair
Data Source
AI summary
A method and a kit for detecting influenza A and B viruses provides a primer group that can simultaneously detect influenza A virus and influenza B virus, in which a first primer pair uses a specific genome fragment of influenza A virus as an amplification target, and a second primer pair uses a specific genome fragment of influenza B virus as an amplification target. The primer group can withstand at least 300 nanograms (ng) of human genomic DNA interference in reverse transcription polymerase chain reaction when detecting influenza A and B viruses, thereby making detection sensitivity less susceptible to interference from human genomic DNA in the sample.