Improvement in production and purification bioprocesses of bacterially expressed anti-alphaIIbbeta3 human single-chain FV antibodies

Remy Robert, A M Noubhani, Marie-Josee Jacobin, X Santarelli, G Clofent-Sanchez

Research output: Contribution to journalArticleResearchpeer-review

8 Citations (Scopus)

Abstract

Production of anti-alphaIIbbeta3 (anti-alphaIIbbeta3)-binding single-chain FV (scFv) fragments obtained from combinatorial libraries of IgG human antibodies is of broad interest for imaging and treatment of acute coronary syndromes. The objective of our work was to design an optimized production of one selected anti-alphaIIbbeta3-binding scFv fragment for subsequent in vivo animal studies. Fed-batch fermentation was initiated with 2TY media supplemented with 0.1 M glucose. This growing batch culture was used as a starting point for further fed-batch induction, in which a media without glucose containing 1 mM IPTG and 0.4 M saccharose was continuously added. Subsequent purification was performed on the whole cell extract in native conditions over an immobilized copper-ion affinity column. The improved conditions allowed the recovery of 5 mg of highly purified scFv fragments as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The bioactivity of the scFv fragments was further monitored by ELISA, cytometric and immunohistochemical methods.
Original languageEnglish
Pages (from-to)43 - 51
Number of pages9
JournalJournal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences
Volume818
Issue number1
DOIs
Publication statusPublished - 2005
Externally publishedYes

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