Alkaline Trypsin Harvesting Solution for Single-Step Virus Recovery
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Solution Overview
Problem
Existing methods for harvesting viruses, such as vaccinia virus, herpes simplex virus, and adenovirus, are inefficient, energy-intensive, and damage viral particles, limiting the production scale and quality of viral biological products.
Innovation Solution
A single-step harvesting method using a solution composed of trypsin, a pH buffer, and optionally a nuclease, with a pH range of 7.5 to 10.5, effectively lysing cells to release viruses without freeze-thaw or mechanical disruption, ensuring viral integrity and activity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If freeze-thaw method is used to lyse cells, then intracellular viruses can be released, but energy consumption increases and production time extends
Solution Approach 1:
The patent changes the chemical parameters of the lysis solution by adjusting pH to alkaline range (7.5-10.5) and optimizing enzyme concentrations (trypsin 0.01-0.12%, nuclease 1-100 IU/ml) to achieve effective cell lysis without requiring energy-intensive freeze-thaw cycles, thereby reducing energy consumption while maintaining high virus yield
2Quantity of substance
If freeze-thaw method is used to lyse cells, then intracellular viruses can be released, but production period extends
Solution Approach 1:
The patent applies preliminary action by pre-optimizing the chemical composition of the lysis solution with specific concentrations of trypsin, nuclease, and pH buffer, allowing cells to be lysed rapidly upon contact without requiring repeated freeze-thaw cycles, thereby significantly shortening the production time while maintaining high virus yield
3Ease of manufacture
If chemical lysis reagents are used to harvest intracellular viruses, then cell lysis is achieved, but viral envelope is destroyed and biological activity is lost
Solution Approach 1:
The patent carefully controls the parameters of chemical reagents by using mild alkaline pH (7.5-10.5) and low concentrations of enzymes (trypsin 0.01-0.12%, nuclease 1-100 IU/ml), which are sufficient to lyse cells but gentle enough to preserve the viral envelope and maintain biological activity, thus resolving the contradiction between lysis efficiency and virus integrity
4Quantity of substance
If multiple harvesting steps are used, then virus extraction is more thorough, but process complexity increases
Solution Approach 1:
The patent merges multiple harvesting steps into a single-step process by formulating a comprehensive lysis solution that contains all necessary components (trypsin, nuclease, pH buffer) to simultaneously achieve cell lysis, virus release, and nucleic acid degradation, thereby simplifying the process while maintaining high virus yield
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 significantly improves virus yield by 5-10 times, simplifies the process, reduces costs, and ensures high-concentration, high-purity viral products suitable for large-scale production.
Implementation Method 1
the harvesting solution composition comprises a trypsin
Implementation Method 2
optionally, a nuclease
Data Source
AI summary
The present invention provides a method for producing a virus and a harvesting solution composition. The method includes culturing cells, wherein the cells have been inoculated with viruses or have been transfected with viral packaging elements; and contacting the cultured cells with a harvesting solution composition to harvest the viruses in a single step, wherein the harvesting solution composition comprises a trypsin, a pH buffer, and, optionally, a nuclease, and wherein the pH of the harvesting solution composition is greater than 7.5 and no more than 10.5. The virus production method of the present invention has advantages of simple operation, easy scale-up, stable yield and so on, and the yield is unexpectedly and significantly improved compared to the prior art, and it can ensure the integrity of the viral particles without damaging the biological activity of the viruses. Therefore, it is very suitable for large-scale production of viruses.