Viral Detection via Universal Primers and Mass Spectrometry
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Solution Overview
Problem
Current methods for detecting adventitious viral contaminants in cell cultures and biologicals are not specific, do not provide information on the source of reverse transcriptase activity, and are not practical for high-throughput, quality-controlled detection of a broad range of viral agents, including unknown or uncharacterized viruses.
Innovation Solution
The use of oligonucleotide primers and kits that hybridize to conserved sequence regions of viral nucleic acids, amplifying specific regions to produce molecular mass and base composition signatures for identification and quantification of adventitious viruses using mass spectrometry analysis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If PCR-based detection methods are used to increase sensitivity and identify particular adventitious agents, then detection sensitivity improves, but the complexity of the testing protocol and time required for analysis increases
Solution Approach 1:
The patent combines multiple detection functions into a single mass spectrometry-based assay. By using universal primers that amplify conserved viral regions followed by mass spectrometric analysis, the method merges sensitivity enhancement, viral identification, and characterization into one integrated protocol, eliminating the need for multiple separate PCR-based tests
Solution Approach 2:
The invention employs universal primers targeting conserved viral sequences that can detect multiple virus families simultaneously. This multi-functional approach allows a single assay to identify diverse adventitious agents including retroviruses, parvoviruses, and other contaminants without requiring virus-specific protocols for each agent
2Reliability
If multiple PCR strategies and co-cultivation assays are used to detect retroviral integration and tumorigenesis risk, then detection comprehensiveness improves, but the time and resources required for complete analysis increase
Solution Approach 1:
The patent performs preliminary amplification of conserved viral regions using universal primers before mass spectrometric analysis. This preliminary action captures potential retroviral sequences early in the workflow, allowing subsequent rapid identification through mass spectrometry without requiring time-consuming follow-up co-cultivation assays or multiple sequential PCRs
3Adaptability or versatility
If broad-range PCR and mass spectrometry analysis are used to detect unknown or uncharacterized viruses, then adaptability to diverse viral agents improves, but the cost of implementation increases
Solution Approach 1:
The patent replaces traditional sequencing-based viral identification with mass spectrometry. This substitution eliminates the need for expensive, time-consuming Sanger sequencing or next-generation sequencing while maintaining broad detection capability. The mass spectrometric analysis of amplified viral DNA provides rapid, cost-effective identification of unknown viruses through molecular weight fingerprinting
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
Enables rapid, sensitive, and cost-effective detection of adventitious viral agents, including previously unknown viruses, at the species or sub-species level, facilitating the identification and quantification of contaminants in a high-throughput, quality-controlled manner.
Implementation Method 1
contacting nucleic acids from the sample with at least one primer pair, wherein each of the two primers of each of said at least one primer pair hybridizes to a conserved sequence region of a Dependovirus nucleic acid
Implementation Method 2
amplifying the two conserved sequence regions and the variable region to produce an amplification product
Implementation Method 3
determining the molecular mass of said amplification product by mass spectrometry
Data Source
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AI summary
The present invention provides oligonucleotide primers, compositions, and kits containing the same for rapid identification of viruses by amplification of a segment of viral nucleic acid followed by molecular mass analysis.