Yuanxing Gene has a genetic recombinant virus purification platform, which can purify various types of viruses, including adenovirus, vaccinia virus, lentivirus, herpes virus, adeno-associated virus, Newcastle disease virus, etc. The purification scale is adapted to the virus culture scale, and the maximum purification of the harvest liquid from a 200L scale bioreactor can be completed, and a recombinant virus product with high viral activity, high purity and high concentration that meets the requirements of drug quality standards can be obtained, which can be used for basic research and preclinical research and clinical studies.
The main operations of purification include clarification, ultrafiltration concentration, and chromatography.
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Clarification
Clarification Clarification of the cell harvest is the first step in downstream purification in a biopharmaceutical process, with the goal of efficiently separating cells, cell debris, and the virus of interest from the harvest, usually using centrifugation, filtration, or tangential flow techniques. Different clarification methods can be selected according to different experimental designs.
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Centrifugal
Use centrifugal force to separate the components of liquid and solid particles or liquid and liquid mixture.
Commonly use high-speed refrigerated centrifuges for primary separation and purification biological products.
Our company uses the Avanti J-26S XP centrifuge to remove large particles of impurities such as cell debris by continuous flow centrifugation or ultracentrifugation to concentrate the virus.
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Filter
Pre-filtration is a common method for removing particulate impurities and reducing bioburden in the biopharmaceutical field. Clarification filtration can effectively remove large particle size particles through the effect of size exclusion and charge adsorption, which can greatly increase the flux of downstream terminal membrane filtration and make the operation of the entire filtration system more economical and reliable.
Minimal loss of active ingredients by filtration, no morphological changes, no chemical changes, large processing scale and high efficiency.
We use disposable filter cartridges or depth filters with non-specific adsorption and high product yield to remove impurities, with high biological safety and no residue between batches.
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Ultrafiltration
Ultrafiltration is a pressurized membrane separation technology, that is, under a certain pressure, small molecular solutes and solvents pass through a special membrane with a certain pore size, while macromolecular solutes cannot pass through and stay on one side of the membrane, so that macromolecular species were partially purified.
The pore size of the ultrafiltration membrane is less than 0.1 μm, and the pore size can be selected according to the needs. It has the separation characteristics of high efficiency and low energy consumption. In virus purification, some small-molecule impurities can be removed, and the virus harvest solution can be concentrated at the same time.
Our company has AKTA flux6 and uniflux 30 ultrafiltration systems. According to different treatment volumes, the corresponding tangential flow ultrafiltration system is selected, which has low shear force, high flux, high product activity recovery rate and large processing capacity.
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Chromatography
Our company has AKTA pure and AKTA pilot purification systems, which can be used for column chromatography purification of different scales.
Our company uses column chromatography to purify the virus, mainly including ion exchange chromatography, gel chromatography, multimodal chromatography. The chromatography process does not produce heat, external force, and has high recovery, high resolution.It can be magnified linearly , automated, and has numerous successful cases.
Ion exchange chromatography is a chromatographic method in which the ion exchanger is used as the stationary phase and the difference in binding force between the component ions in the mobile phase and the counter ions on the exchanger is reversibly exchanged. In adenovirus purification, according to different virus types, a variety of resins such as Source 30Q, Q Sepharose XL, Fractogel DEAE, Capto Q, etc. were applied respectively.
Gel chromatography is a technique for separating proteins according to their molecular weight, also known as molecular sieve filtration, size exclusion chromatography, etc. Its outstanding advantages are that the gel used in chromatography is an inert carrier, has no charge, weak adsorption, mild operating conditions, does not require organic solvents, and has a unique ability to maintain the physical and chemical properties of the separated components. It has a good separation effect for high molecular substances.
Multi-mode chromatography refers to a chromatography method with dual functions of size separation and adsorption chromatography. A typical multimodal chromatography resin, Capto Core 700, is designed for purification of viruses and other biological macromolecules. Capto Core 700 consists of an activated ligand core and an inert shell. The inert shell can exclude macromolecules (exclusion molecular weight ~ Mr, 700000 [700kDa]), prevent them from entering the core through the pores on the shell. While these macromolecules are collected in the column flow-through, smaller impurities bind to the octylamine ligand within the microspheres. These features make Capto Core 700 an excellent alternative to size exclusion chromatography resin typically used in the final stages of virus purification in vaccine production.
