Recombinant human nerve growth factor purifying method based on CHO cell expression system

Abstract

The invention relates to a recombinant human nerve growth factor purifying method based on a CHO cell expression system. The recombinant human nerve growth factor purifying method based on the CHO cell expression system comprises the following steps: 1), centrifugating supernatant, successively removing living cells and cell fragments, and removing impurities and bacteria by filtering; 2), applying a supernatant sample to an EMD SO3-(M) chromatography column, rinsing unbound protein and impure protein, eluting and collecting a target protein peak; 3), concentrating the protein with ultrafiltration equipment with a molecular weight cutoff of 3K; and 4), applying the sample to Superdex 75-containing chromatography column and collecting the target protein peak, namely a recombinant human nerve growth factor stoste. According to the CHO cell expression system and the recombinant human nerve growth factor purifying method based on the CHO cell expression system, the recombinant human nerve growth factor with the purity greater than 98% and the specific activity higher than the 5* 10 to the power of 5 AU / mg can be prepared; compared with other methods, the recombinant human nerve growth factor purifying method based on the CHO cell expression system has the advantages that the steps are reduced, the production cycle is greatly shortened, and the processing capacity and the yield are improved; therefore, the recombinant human nerve growth factor purifying method based on the CHO cell expression system is application for large-scale production.

Description

technical field [0001] The invention relates to the technical field of biomedicine extraction, in particular to a method for extracting recombinant human nerve growth factor from the expression supernatant of CHO cells. Background technique [0002] Human Nerve Growth Factor (hNGF) is the earliest discovered important nutritional factor in the nervous system. It plays an important role in the growth, development, differentiation and regeneration of central and peripheral nerve cells. The survival and differentiation of these neurons plays an important role in protecting the normal function of the nervous system; and hNGF has important clinical value in the treatment of brain injury, senile dementia, brain Parkinson's syndrome and other diseases related to nerve damage and neurodegeneration (Lv Xianfeng, Liu Zhiqiang, Wang Dong, et al., A method for purifying nerve growth factor from cobra venom suitable for large-scale production, application publication number: CN101845093A...

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Problems solved by technology

[0004] However, most of the traditional purification processes have cumbersome steps and relatively backward chromatography packing materials, which have disadvantages such as low yield, long time-consuming, high probability of protein contamination, etc.

Due to the relatively expensive cost of the CHO cell expression system, if the traditional purification process is used, there will be huge disadvantages in terms of protein yield, activity and time loss, thereby increasing product costs, which has always affected the industrialization of CHO cell expression of recombinant human nerve growth factor. main reason for production

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