Method for producing monoclonal antibody by using hybridoma and produced monoclonal antibody

Abstract

The invention discloses a method for producing a monoclonal antibody by using hybridoma. The method comprises the following steps: placing resuscitated hybridoma in a first cell culture medium for performing sub-culture and enlarge culture, wherein the volume fraction of serum in the first cell culture medium is larger than 10 percent; transferring the hybridoma into a second cell culture medium for performing enlarge culture, wherein the volume fraction of serum in the second cell culture medium is less than or equal to 10 percent; transferring the hybridoma into a serum-free cell culture medium for culturing, and performing monoclonal antibody production. By adopting the method, the first cell culture medium is used for performing eutrophic culture, and the second cell culture medium isused for performing uniform group culture, so that the concentration and volume of hybridoma of different batches can keep consistent at the start of antibody production, the whole production flow canbe widely applied to different hybridoma plants, and the growth period of the antibody is stable. The monoclonal antibody produced in the serum-free cell culture medium has high product quality.

Description

technical field [0001] The invention belongs to the technical field of hybridoma cell culture and monoclonal antibody production, and in particular relates to a method for producing monoclonal antibody by using hybridoma cells and the produced monoclonal antibody. Background technique [0002] Hybridoma cells (hybridoma) are cells fused with myeloma cells and monoclonal B cells. The fusion cells not only have the ability of myeloma cells to replicate infinitely, but also have the ability of mature monoclonal B plasma cells to continuously secrete monoclonal antibodies. Features. In the industrial production of antibodies, such fusion cells are used for large-scale production of monoclonal antibodies, and such fusion cells are called hybridoma cells. [0003] The traditional production scheme of using hybridoma cells to produce monoclonal antibodies is generally to culture hybridoma cells in RPMI medium containing 10% fetal bovine serum, and the purity of monoclonal antibodi...

AI Technical Summary

Problems solved by technology

[0005] The disadvantage of this production scheme is that (a) hybridoma cells complete the production of monoclonal antibodies in 2% fetal bovine serum RPMI culture fluid, but there are still more proteins in fetal bovine serum, which directly leads to the production of hybridoma cells. The purity of the secreted antibody is not high, resulting in poor purity (about 90%) and quality of subsequent products; (b) the entire production cycle takes 25 days, which is long; (c) due to the low nutrient content of the medium, more species are hybridized Tumor cells cannot complete expansion culture in 2% low serum RPMI medium

[0007] The weak point of this production scheme is: (a) the hybridoma cells have a 15-day fetal calf serum RPMI culture solution serum concentration gradual decline culture process before being transferred to the serum-free medium, which causes the production cycle to be extended for 15 days, and the whole The antibody production cycle needs at least 35 days; (b) During the serum gradient culture process, it is difficult for the hybridoma cells to maintain the best growth state, the quality of antibody production is worrying, and the human operation will lead to the instability of the cell culture state between batches, and finally It will lead to unstable quality of antibody products between batches; (c) Different hybridoma cells have different adaptability to the serum gradual decline process, resulting in large differences in the production cycle of different types of hybridoma cells. The process cannot be consistent; (d) the hybridoma cells complete the production of monoclonal antibodies in 2% fetal bovine serum RPMI culture medium, but there are still many proteins in the fetal bovine serum, which directly leads to the purity of the antibodies secreted by the hybridoma cells Not high, causing follow-up product purity (about 90%) and quality to be still relatively poor

However, the development of serum-free medium has not yet achieved a breakthrough, and the quality and yield of monoclonal antibodies produced are still not high