Chemical component limited culture medium for HEK293 cell culture and replication and amplification of adenovirus and adeno-associated virus

A cell culture and chemical composition technology, applied in the direction of animal cells, vertebrate cells, artificial cell constructs, etc., can solve the harsh transportation and storage conditions of culture medium products, inconvenient transportation and storage, inconvenient solution for customers, and insufficient product titer, etc. problems, to achieve the effect of ensuring cell viability and expansion speed, high product safety and stability, and loose transportation and storage conditions

Pending Publication Date: 2022-08-02
上海东富龙生物试剂有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] 1. The formula contains proteins, protein hydrolyzates and other substances that are not chemically limited, which makes the final product less safe;
[0008] 2. The medium with non-chemically limited components has insufficient stability, and it is difficult to control the batch-to-batch difference during the large-scale production of the medium;
[0009] 3. The non-chemically restricted substances contained are sensitive to the environment, resulting in harsh conditions for the transportation and storage of culture medium products;
[0010] 4. Substances such as protein or protein hydrolyzate affect the separation and purification of the final product, and are difficult to remove in the downstream separation and purification process. The requirements for separation and purification are high, the cleaning of the purification column is difficult, the separation and purification time is prolonged, and the service life of the purification column is reduced;
[0011] 5. Full formula grinding cannot effectively guarantee product quality. Most products contain multiple components such as basal medium and additives, which cause inconvenience to product transportation and storage and customer liquid preparation;
[0012] 6. The medium product with limited chemical composition is single, and there is no corresponding development for the R&D and production of adenovirus and adeno-associated virus. After virus transfection, the activity and continuous amplification ability of cells decrease, resulting in insufficient product titer

Method used

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  • Chemical component limited culture medium for HEK293 cell culture and replication and amplification of adenovirus and adeno-associated virus
  • Chemical component limited culture medium for HEK293 cell culture and replication and amplification of adenovirus and adeno-associated virus
  • Chemical component limited culture medium for HEK293 cell culture and replication and amplification of adenovirus and adeno-associated virus

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Experimental program
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Embodiment 1

[0072] The components of the chemical composition-limited culture medium of the present embodiment and their contents are as follows:

[0073] The composition and content of amino acids are:

[0074]

[0075] The components of the vitamin and its content are:

[0076]

[0077] The composition and content of trace elements are:

[0078]

[0079]

[0080] The components and contents of inorganic salts are:

[0081]

[0082] The composition and content of fatty acids are:

[0083] Linoleic acid 0.5mg / L

[0084] Linolenic acid 0.3mg / L.

[0085] Prepare the raw material components according to the above specific contents, fully dissolve them in sterile deionized water, adjust the pH to 6.9-7.0, adjust the volume to 10L, filter with a filter membrane with a pore size of 0.22 μm, and store at 4°C for later use (hereinafter referred to as CDM medium).

[0086] The cells used in this example were HEK293 cells, which were purchased from CTCC (Mason cells) in China. ...

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Abstract

The invention discloses a chemical component limited culture medium for HEK293 cell culture and replication and amplification production of adenovirus and adeno-associated virus. The chemical component limited culture medium consists of amino acid, vitamins, trace elements, inorganic salt and fatty acid. The chemical components of the culture medium are limited, the culture medium is clear in chemical element components and free of serum and protein, plant hydrolysate is not added, small molecular compounds with known structures and functions are adopted for substitution, HEK293 cell culture metabolism and replication and amplification of adenoviruses and adeno-associated viruses are more facilitated, and the culture medium is safer and more effective.

Description

technical field [0001] The invention belongs to the field of adenovirus, adeno-associated virus and bioengineering, and particularly relates to a chemical-component-limited medium suitable for HEK293 suspension large-scale amplifying culture and adenovirus and adeno-associated virus replication and amplification production. Background technique [0002] Adenovirus and adeno-associated virus (AAV) are two different types of viral vectors commonly used for gene delivery. These two recombinant virus systems have obvious advantages in infecting the host, such as low immunogenicity and infecting dividing and non-dividing cells. The differences between the two are mainly in the packaging capacity and level, the onset and duration of gene replication and amplification, and the difference in immune response. Adenoviruses have a capacity of about 8.5 kilobases and can replicate high levels of proteins and transient genes. The packaging capacity of AAV is about 4.5 kilobases, the le...

Claims

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Application Information

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IPC IPC(8): C12N5/071
CPCC12N5/0686C12N2500/32C12N2500/38C12N2500/05C12N2500/36
Inventor 胥坤杨茜茜
Owner 上海东富龙生物试剂有限公司
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