Nanopore Viral Particle Quantification and Infectivity Assessment
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Solution Overview
Problem
Current methods for quantifying viral particles are indirect, variable, and lack precision, particularly in measuring infectivity and replication efficiency, which hampers gene therapy and vaccine development by providing inaccurate composition analysis of viral vector products.
Innovation Solution
A system combining nanopore technology to discriminate viral particles based on biophysical properties and single-molecule RNA fluorescence in situ hybridization (smFISH) for precise counting and infectivity assessment, enabling direct quantification of viral particles and their functional titer, integrated into standard laboratory settings without requiring large equipment.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Ease of operation
If indirect measurement methods (qRT-PCR, ELISA, plaque assays) are used to quantify viral particles, then the measurement process is simplified and can be performed with standard equipment, but the measurement precision and accuracy of total particle count is compromised
Solution Approach 1:
The patent replaces indirect biochemical measurement systems (qRT-PCR, ELISA, plaque assays) with direct physical measurement using nanopore technology. The nanopore system measures electrical current blockage caused by individual viral particles passing through the pore, providing direct physical counting rather than indirect biochemical inference. This substitution enables single-particle resolution and accurate total particle quantification while maintaining operational simplicity.
Solution Approach 2:
The patent changes the measurement parameter from biochemical indicators (genome copy number, antigen presence, plaque formation) to direct physical parameters (electrical current blockage, particle size, charge). By measuring the electrical signature and physical dimensions of individual particles as they translocate through the nanopore, the system achieves direct counting of total particles independent of infectivity or genome presence, resolving the contradiction between ease of measurement and precision.
2Device complexity
If existing quantification methods are used, then the assay can be performed with standard laboratory equipment, but the ability to measure infectivity and replication efficiency simultaneously with total particle count is lost
Solution Approach 1:
The patent merges multiple measurement capabilities into a single nanopore system. The system simultaneously measures total particle count, particle size distribution, charge characteristics, and when combined with smFISH, infectivity and replication efficiency. This consolidation allows comprehensive viral characterization without requiring separate assays, large equipment, or sacrificing any measurement capability.
Solution Approach 2:
The nanopore system serves multiple functions: it counts total particles, characterizes particle size and charge, and when integrated with smFISH, assesses infectivity and replication efficiency. This multi-functionality is achieved within a single device that can be implemented in standard laboratory settings, eliminating the need for multiple specialized assays and equipment while preserving all measurement information.
3Productivity
If batch-to-batch variability in virions is not accounted for, then the viral product composition analysis is faster and simpler, but the reliability of viral delivery assays decreases
Solution Approach 1:
The patent implements comprehensive feedback by measuring multiple parameters (total particle count, size distribution, charge, infectivity, replication efficiency) that characterize the complete viral product composition. This detailed feedback on batch-to-batch variability enables precise quality control and assay optimization, improving reliability while maintaining productivity through the efficiency of the nanopore measurement system.
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 allows for accurate, precise measurement of viral particle counts and infectivity, optimizing virus generation for maximum infectious payload and enhancing the robustness and quality of viral delivery assays, as well as providing a standard for comparing viral quantification across experiments and laboratories.
Implementation Method 1
electrodes configured to energize and apply a voltage gradient across the nanopore between the interior cavity and exterior cavity, the voltage gradient of sufficient magnitude to induce particles to migrate via the nanopore
Implementation Method 2
produce an electronic signature representative of at least one property of the particles
Data Source
AI summary
A system and method for assessing properties of particles, the system comprising: a first structure defining an interior cavity, configured to contain an interior cavity fluid, and a barrier separating the interior cavity from an exterior cavity configured to contain an exterior cavity fluid, the barrier defining a nanopore therethrough fluidically coupling the interior and exterior cavities, the first structure further comprising at least one feature configured to interface with at least one complementary feature of a second structure that defines the exterior cavity; and electrodes configured to energize and apply a voltage gradient across the nanopore between the interior cavity and exterior cavity, the voltage gradient of sufficient magnitude to induce particles to migrate via the nanopore between the interior cavity and the exterior cavity and produce an electronic signature representative of at least one property of the particles.