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(Reference retrieved automatically from Web of Science through information on FAPESP grant and its corresponding number as mentioned in the publication by the authors.)

Laboratory Production of Human Prolactin from CHO Cells Adapted to Serum-Free Suspension Culture

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Author(s):
Arthuso, Fernanda Santos [1] ; Bartolini, Paolo [1] ; Jorge Soares, Carlos Roberto [1]
Total Authors: 3
Affiliation:
[1] Inst Pesquisas Energet & Nucl IPEN CNEN SP, Ctr Biotechnol, BR-05508900 Sao Paulo - Brazil
Total Affiliations: 1
Document type: Journal article
Source: Applied Biochemistry and Biotechnology; v. 167, n. 8, p. 2212-2224, AUG 2012.
Web of Science Citations: 5
Abstract

Human prolactin (hPRL) is a polypeptide with 199 amino acids and a molecular mass of 23 kDa. Previously, a eukaryotic hPRL expression vector was used to transfect Chinese hamster ovary (CHO) cells: this work describes a fast and practical laboratory adaptation of these transfected cells, in similar to 40 days, to grow in suspension in serum-free medium. High cell densities of up to 4.0 x 10(6) cell/ml were obtained from spinner flask cultures and a stable and continuous production process was developed for at least 30 days. Two harvesting strategies were set up, 50 or 100 % of the total conditioned medium being collected daily and replaced by fresh culture medium. The volumetric productivity was 5-7 mu g hPRL/ml, as determined directly in the collected medium via reversed-phase HPLC (RP-HPLC). A two-step process based on a cationic exchanger followed by size exclusion chromatography was applied to obtain purified hPRL from conditioned medium. Two hPRL isoforms, non-glycosylated and glycosylated, could also be separated by high-performance size-exclusion chromatography (HPSEC) and, when analyzed by RP-HPLC, HPSEC, Western blotting, and bioassay, were found to be comparable to the World Health Organization International Reference Reagent of hPRL. These results are useful for the practical scale-up to the pilot and industrial scale of a bioprocess based on CHO cell culture. (AU)