Recombination fibroblast preparation

The Western blot method is often used in the analysis of host cell impurities. It can be used to identify a recurring impurity. O Keefe et al. used a Western blot to identify an E. coli protein impurity in the preparation of the recombinant fibroblast growth factor (aFGF).29 By using specific antisera to the E. coli host cell proteins, they were able to isolate the impurity and determine its N-terminus amino acid sequence to confirm its identity. Antibodies could be used to determine the concentration of this impurity in sample preparations. 

Several of these IFNs of mouse and human lymphocytes and fibroblasts are available commercially and have been best prepared in quantity by recombinant DNA procedures because they are produced in very small amounts by the cells. The commercial materials do not generally require further purification for their intended purposes. [Pestkas, Interferons and Interferon standards and general abbreviations. Methods Enzymol, Wiley Sons, 119 1986, ISBN 012182019X Lengyel, Biochemistry of interferons and their actions, Ann Bev Biochem 51 251-282 7982 De Maeyer and De Maeyer-Guignard, Interferons in The Cytokine Handbook, 3rd Edn, Thomson et al. Eds, pp. 491-516 7998 Academic Press, San Diego, ISBN 0126896623.]... 

Classification Human interferons have been classified into three type s, a, p, and y, on the basis of their antigenicity. The a interferons are primarily leukocytic, whereas the p and y interferons are produced by connective tissue fibroblasts and T lymphocytes, respectively. Recombinant DNA techniques in bacteria have made available large quantities of a interferon/species A—designated a2(A) interferon—which has the advantage of being purer than previous preparations derived from leukocytes. Currently the other interferons have also been obtained by recombinant techniques. 

Interferon beta is used in the form of natural fibroblast or recombinant preparations (interferon beta-la and interferon beta-lb) and exerts antiviral and antiproliferative properties similar to those of interferon alfa. Although its efficacy has been debated (1), interferon beta has been approved for the treatment of relapsing-remitting multiple sclerosis, and more recently for secondary progressive multiple sclerosis. 

These studies can be done with liver microsomes, hepatocytes, and recombinant preparations of CYP enzymes. Microsomes are membrane fragments prepared from homogenized liver, while hepatocytes are liver cells that have been collected from collagenase-perfused liver. These are then cocultured with collagen, and fibroblasts... 

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