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Author (up) Rosa, P.A.J.; Azevedo, A.M.; Aires-Barros, M.R. doi  openurl
  Title Application of central composite design to the optimisation of aqueous two-phase extraction of human antibodies Type Journal Article
  Year 2007 Publication Journal of Chromatography. A Abbreviated Journal J Chromatogr A  
  Volume 1141 Issue 1 Pages 50-60  
  Keywords Analysis of Variance; Animals; Antibodies/*chemistry/*isolation & purification; Buffers; Chemical Fractionation/*methods; Horses; Humans; Hydrophobicity; Isoelectric Point; Models, Biological; Molecular Weight; Myoglobin/chemistry/isolation & purification; Osmolar Concentration; Phase Transition; Polyethylene Glycols; Serum Albumin/chemistry/isolation & purification; Sodium Chloride  
  Abstract The partition of human antibodies in aqueous two-phase systems (ATPSs) of polyethylene glycol (PEG) and phosphate was systematically studied using first pure proteins systems and then an artificial mixture of proteins containing 1mg/ml human immunoglobulin G (IgG), 10mg/ml serum albumin and 2mg/ml myoglobin. Preliminary results obtained using pure proteins systems indicated that the PEG molecular weight and concentration, the pH value and the salts concentration had a pronounced effect on the partitioning behaviour of all proteins. For high ionic strengths and pH values higher than the isoelectric point (pI) of the contaminant proteins, IgG could be selectively recovered on the top phase. According to these results, a face centred composite design was performed in order to optimise the purification of IgG from the mixture of proteins. The optimal conditions for the isolation of IgG were observed for high concentrations of NaCl and low concentrations of both phase forming components. The best purification was achieved using an ATPS containing 8% (w/w) PEG 3350, 10% (w/w) phosphate pH 6 and 15% (w/w) NaCl. A recovery yield of 101+/-7%, a purity of 99+/-0% and a yield of native IgG of 97+/-4% were obtained. Back extraction studies of IgG to a new phosphate phase were performed and higher yields were obtained using 10% phosphate buffer at pH 6. The total extraction yield was 76% and the purity 100%.  
  Address IBB Institute for Biotechnology and Bioengineering, Centre for Biological and Chemical Engineering, Instituto Superior Tecnico, Av Rovisco Pais, 1049-001 Lisbon, Portugal  
  Corporate Author Thesis  
  Publisher Place of Publication Editor  
  Language English Summary Language Original Title  
  Series Editor Series Title Abbreviated Series Title  
  Series Volume Series Issue Edition  
  ISSN 0021-9673 ISBN Medium  
  Area Expedition Conference  
  Notes PMID:17196214 Approved no  
  Call Number Serial 1842  
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