Serum-free medium for MDCK cell large-scale adherent culture and single-cell suspension culture

Abstract

The invention relates to the culture medium research and development technical field of modern biological technology and provides a serum-free medium for MDCK cell large-scale adherent culture and single-cell suspension culture, which comprises 21 amino acids, 6 vitamins, 8 salts, 8 lipids, 4 trace elements, 2 buffers, 1 protein hydrolysate, 1 acid-base indicator and 6 other additives. The serum-free medium can be prepared by the conventional preparation method, and an application method thereof is the conventional method. The serum-free medium has the beneficial effects that: the serum-free medium does not contain serum, has clear components, is beneficial for separating and purifying the product and improves the product quality; the serum-free medium supports long-term subculture of MDCK cells and does not require long-term and complex adaptation process; and the serum-free medium can well support the adherent growth and single-cell suspension growth of the MDCK cells, has clear components and easy preparation and utilization, and is suitable for mass production of biological products.

Description

【Technical field】 [0001] The invention relates to the technical field of medium research and development of modern biotechnology, in particular to a serum-free medium suitable for large-scale adherent culture of MDCK cells and single cell suspension culture. 【technical background】 [0002] Preventing viruses from endangering human beings is a matter of great concern to all countries. For many years, human beings have suffered huge losses every year due to viruses, such as rabies and bird flu. Rabies is caused by the rabies virus and is one of the most serious infectious diseases. Once the onset occurs, the mortality rate is very high, and so far there is no special and effective therapeutic drug except the vaccine for prevention. In addition, the harm caused by influenza virus is also very great. Every year, 5-15% of people are infected by seasonal influenza, 3-5 million severe cases, and up to 500,000 people die from it. What's more serious is that the current influenza v...

AI Technical Summary

Problems solved by technology

[0006] However, the traditional MDCK cell vaccine production process requires the use of serum

The use of serum has several obvious disadvantages: firstly, due to different animal individuals, different places of origin and batch numbers of serum, the quality of serum batches varies greatly, making it difficult to standardize experiments and production; secondly, the composition of serum is complex, which contains both The components of cell growth also contain components that inhibit the growth of other cells, and also contain substances that are toxic to cells, such as complements, antibodies, bacterial toxins, etc., which will affect the growth of cells and even cause cell death; again, in serum Mycoplasma and viruses may be introduced into the samples, which may have a potential impact on the cells, which may lead to experimental failure or unreliable experimental results; fourth, when dealing with large-scale production, the source of serum is becoming more and more difficult and expensive; finally, The introduction of serum will also increase the difficulty of downstream separation and purification of products, increase production costs, make it difficult to achieve standardized production, and may bring potential safety hazards

SAFC Biosciences, a subsidiary of Sigma-Aldrich in the United States, launched a serum-free and animal protein-free commercial medium Ex-Cell MDCK in 2006, and claimed that the medium can support MDCK cell attachment and single-cell suspension growth, but until At present, there is no literature report on the cultivation of MDCK cells in single cell suspension in this medium

Although these commercial media can support the stable growth of cells and be used in the production of biological products, they also have some obvious disadvantages, such as: the composition is kept secret, the chemical composition is not clear, which brings difficulties to the scientific research and production process; the price is relatively expensive, It is only suitable for some small-scale experimental research and not suitable for large-scale production; it can only support the adherent growth of MDCK cells. In the process of large-scale culture, cell-attached substrates such as microcarriers are often used, which not only increases the cost but also hinders the cost of biological products. Post-purification brings trouble

Images