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A kind of separation and purification method of rhtrail thalline

A technology for separation and purification of bacterial cells, applied in the field of separation and purification of rhTRAIL bacterial cells, can solve problems such as being unsuitable for large-scale industrial production, high production cost, long working cycle, etc., and achieve effective quality control, high production efficiency, and convenience. Effects of operation and maintenance

Active Publication Date: 2016-08-03
深圳未名新鹏生物医药有限公司
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Problems solved by technology

[0003] Although the original purification process can finally obtain samples that meet the quality requirements, it is not suitable for the needs of large-scale industrial production due to the long working cycle, low recovery rate, and excessive production costs.

Method used

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  • A kind of separation and purification method of rhtrail thalline

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Experimental program
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Embodiment

[0060] see figure 1 , according to the following steps, the sample rhTRAIL cells were isolated and purified:

[0061] Step 1: Crude Pure:

[0062] 1.1 High pressure homogenate crushing:

[0063] Weigh a certain amount of cryopreserved rhTRAIL cells and add an appropriate amount of B1 solution, mash them evenly with a tissue grinder until there are no lumps of cells, add an appropriate amount of B1 solution again to make the final cell concentration 10%, and mash The homogeneous bacterial suspension was crushed once under a pressure of 1500 bar, and the homogenized crushed liquid was collected and stored in a refrigerator at 4-8°C for later use.

[0064] The B1 solution is 50mM / LTris-HCl, buffer solution with pH 8.5, and 0.1% DTT has been added according to the amount of bacteria before use; it is prepared with water for injection, and is sterilized and filtered with a 0.22um filter membrane after preparation.

[0065] 1.2 High-speed centrifugal separation

[0066] Centrifu...

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Abstract

The invention provides a separation purification method for rhTRAIL thalline, and the separation purification method comprises a crude purification technology and a fine purification technology. The crude purification technology comprises: performing amplified culture and fermentation on rhTRAIL thalline obtained by a recombinant DNA technology, collecting thalline, performing high-pressure homogenate, fragmentation and microfiltration. The fine purification technology comprises: performing ion chromatography, hydrophobic chromatography, gel filtering chromatography and concentration on the crudely pure sample obtained in the crude purification technology to obtain a sample accord with quality requirement. The separation purification method has the advantages of being short in operation period, high in recovery rate and relatively low in production cost, and is applicable to large-scale industrial production.

Description

technical field [0001] The invention relates to the field of separation and purification of biological products, in particular to a method for separation and purification of rhTRAIL cells. Background technique [0002] Tumor necrosis factor-related apoptosis-inducing ligand (TNF-relatedapoptosis-inducingligand, TRAIL) or apoptin 2 ligand (Apo2ligand, Apo2L) is a new member of the TNF family discovered in 1995. The soluble form of TRAIL can induce apoptosis in a variety of tumor cells both in vivo and in vitro. The full-length human TRAIL molecule consists of 281 amino acids, and is a type II transmembrane protein, including the cytoplasmic region, transmembrane region and extracellular region, with a molecular weight of 32.5kD and an isoelectric point of 7.63. The 114th to 281st amino acids of the TRAIL molecule are soluble peptides, which can form the most active homotrimeric structure through a zinc ion. TRAIL exerts its apoptosis-inducing effect by binding to its cell m...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K14/525C07K1/36C07K1/34C07K1/18C07K1/20C07K1/16
CPCC07K14/70575
Inventor 王军张静李靖文
Owner 深圳未名新鹏生物医药有限公司
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