Unlock instant, AI-driven research and patent intelligence for your innovation.

Method for production of factor viii

A technology of factors and viability, applied in the field of production of factor VIII polypeptides

Inactive Publication Date: 2014-07-23
NOVO NORDISK AS
View PDF3 Cites 1 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this harsh treatment does not allow the subsequent reuse of the cells
[0005]Despite understanding of Factor VIII regulation, production of Factor VIII in heterologous systems used in commercial manufacturing is significantly lower than that of other recombinant proteins

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for production of factor viii
  • Method for production of factor viii
  • Method for production of factor viii

Examples

Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0106] binding assay

[0107] 　　 method

[0108] by using 125 Homologous competition assay studies of I-BDD-FVIII and untagged BDD-FVIII Purified B-domain-deleted Factor VIII (BDD-FVIII) (kindly provided by J. Karlsson and L. Thim, Novo Nordisk A / S) Affinity to the cell membrane of HEK293 cells. Cells were washed once in PBS + 1% BSA. Will 5x10 5 The cells were dispensed into microtiter wells, and the plates were cooled to 4°C. During the blocking of the cell surface, the binding of BDD-FVIII was detected. At 4°C, with Annexin V (0.5 μM, Sigma), orthophospho-L-serine (20 mM, Sigma), heparin (100 μg mL-1, Leo Pharmaceuticals) and receptor-associated protein (RAP) 0.5 μM (H.H. Petersen, a gift from Novo Nordisk A / S) was added simultaneously with a constant concentration of 125 I-FVIII (0.5 nM) to prevent endocytosis.

[0109] Plates were incubated for 2 hours at 4°C with gentle shaking. After centrifugation, remove unbound (non-membrane-associated) 125 I-FVIII...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The present invention relates to methods of producing a Factor VIII polypeptide in mammalian cell cultures.

Description

field of invention [0001] The present invention relates to methods for producing Factor VIII polypeptides. Background of the invention [0002] Factor VIII is an important coagulation factor. Mutations in the Factor VIII gene that result in reduced or defective Factor VIII protein cause the genetic disease hemophilia A, which is characterized by recurrent bleeding episodes. Treatment of hemophilia A requires intravenous infusion of plasma-derived or recombinant Factor VIII. [0003] Although plasma-derived factor VIII can be used to treat hemophilia, this approach has several problems, including transmission of the virus to the patient. Therefore, it is preferred to administer recombinantly expressed Factor VIII. [0004] It is difficult to obtain large amounts of Factor VIII from cell culture. Factor VIII is known to be expressed at very low levels in mammalian cells. Factor VIII is also known to be an unstable protein in serum-free or protein-free media. The additio...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/755
CPCC07K14/755
Inventor L.B.约翰逊M.P.科林德P.L.诺拜
Owner NOVO NORDISK AS