Method for separating and purifying human antithrombin-III from human plasma

Abstract

The invention discloses a method for separating and purifying human antithrombin-III from human cryoprecipitate-reduced plasma. The method comprises the operating steps of performing chromatography on the human cryoprecipitate-reduced plasma, performing S / D (solvent / detergent) virus inactivation, and performing affinity chromatography for the second time; performing ultrafiltration; performing sterilizing filtration; filtering with a 20nm nano-filtration membrane to remove viruses; subpackaging; performing freeze-drying; performing dry heat virus inactivation; separating and purifying human antithrombin-III from the human cryoprecipitate-reduced plasma; performing pre-filtration treatment, and then directly separating and purifying antithrombin-III (AT-III) out of the plasma by adopting an affinity chromatography technology; adding an appropriate amount of a protective agent, replacing pasteurella virus inactivation process with an S / D virus inactivation process to reduce the loss and activity and shorten the preparation time; then performing affinity chromatography for the second time to further purify the AT-III, and finally performing the steps of ultrafiltration, virus removal through nano-filtration with a 20nm nano-membrane, freeze-drying and the like, wherein the purity of the final product is more than 95%, the specific activity is more than 6IU / mg, the virus safety is high, and blood plasma can not be influenced during preparation to produce a downstream product.

Description

technical field [0001] The invention belongs to the field of biopharmaceuticals and relates to a method for separating and purifying human antithrombin-III from human cryoprecipitate plasma. Background technique [0002] Human antithrombin-Ⅲ (Antithrombin-Ⅲ, referred to as AT-Ⅲ) is a single peptide chain α-glycoprotein with a molecular weight of about 58kD (kilodaltons), consisting of 423 amino acids, and is a multifunctional serine protease inhibitor , the anticoagulant effect in human plasma accounts for about 70% of the entire anticoagulant effect, and its half-life is 2.69 days, and the half-life is shortened when sick. Degree of acquired lack. AT-Ⅲ products are mainly used to treat congenital hereditary antithrombin-Ⅲ deficiency. AT-Ⅲ deficiency is an autosomal dominant inheritance. According to statistics, the incidence rate of this disease in Europe and the United States is 0.2% to 0.5%. AT-Ⅲ products are also used to treat acquired antithrombin Ⅲ deficiency, which...

AI Technical Summary

Problems solved by technology

[0003] In the 1960s and 1970s, people began to report the process of purifying AT-Ⅲ, but it was limited to laboratory development

In 1972, Heimburger et al first isolated and identified AT-Ⅲ as a2 single-chain glycoprotein, but its recovery rate was low, about 2%.