Virus Titer Quantification via Image Cytometry
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Solution Overview
Problem
Traditional methods for determining viral titer, such as viral plaque assays and ELISA, are slow, labor-intensive, and prone to high inter-assay variability, while flow cytometry instruments are costly and complex, limiting their widespread use for rapid and accurate quantification of infectious viral particles.
Innovation Solution
A high throughput method using serial dilutions of a virus sample, infecting host cells, reacting viral antigens with fluorescent-tagged antibodies, and determining the number of infected cells via image cytometry, such as CellInsight, to plot a linear regression and calculate virus titer in a multiple-well plate format.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Productivity
If traditional viral plaque assays or ELISA are used for virus quantification, then the method is simpler and less expensive, but the analysis time is long and labor intensity is high
Solution Approach 1:
The patent replaces manual mechanical counting methods (plaque assays, ELISA) with automated image cytometry. The system automatically captures images of infected cells, detects fluorescent signals, counts infected cells, and calculates virus titers through software algorithms, eliminating manual labor and accelerating the analysis process significantly
Solution Approach 2:
The patent changes the detection parameter from indirect protein detection (ELISA) or plaque formation (mechanical counting) to direct fluorescent signal detection from individually infected cells. This parameter change enables rapid quantification by counting fluorescent cells rather than measuring protein concentrations or counting plaques manually
2Productivity
If flow cytometry is used for virus quantification, then the analysis speed improves, but the instrument cost and complexity increase
Solution Approach 1:
The patent uses disposable multi-well plates with individual cell compartments instead of expensive, complex flow cytometers. Each well contains a small volume of sample and infected cells, and the entire plate can be discarded after use, eliminating the need for costly instrument maintenance and reducing barriers to adoption
Solution Approach 2:
The patent segments the sample into individual wells, each containing a small volume of virus sample and host cells. This segmentation allows parallel processing of multiple samples simultaneously in a multi-well plate format, achieving high throughput without requiring complex instrumentation - each well can be independently imaged and analyzed
3Measurement precision
If traditional virus quantification methods are used, then the procedure is simpler, but the accuracy and reliability are compromised due to high inter-assay variability
Solution Approach 1:
The system performs self-calibration and internal controls by including reference samples and automated quality checks within each assay run. The image cytometer automatically adjusts for variations in lighting, focus, and detection sensitivity, and includes built-in controls to monitor assay performance, ensuring consistent and reliable results without requiring external calibration
Solution Approach 2:
The patent incorporates feedback mechanisms where the system continuously monitors assay progress and automatically adjusts parameters. Quality control samples provide feedback on assay performance, and the software algorithms automatically correct for detected variations, ensuring high precision and reliability across multiple assays and operators
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 method enables rapid and accurate quantification of infectious viral particles, reducing the need for extensive dilutions and duplicates, allowing for frequent monitoring and optimal harvesting of virus yields for vaccine development and production.
Implementation Method 1
reacting an antigen expressed by the virus in infected cells with an antibody labeled with a fluorescent tag
Data Source
AI summary
This invention relates to high throughput methods of determining a viral titer. The instant invention addresses the need for a more rapid and cost effective method of quantitating infectious viral particles in a sample.