Malpiedi, Luciana P.
Nerli, Bibiana B.
Taqueda, Maria E. S.
Abdalla, Dulcineia S. P.
Pessoa, Jr., Adalberto
Total Authors: 5
 Univ Nacl Rosario, Inst Proc Biotecnol & Quim IPROBYQ, Dept Quim Fis, RA-2000 Rosario, Santa Fe - Argentina
 Univ Sao Paulo, Polytech Sch, Dept Chem Engn, BR-05508000 Sao Paulo - Brazil
 Univ Sao Paulo, Dept Clin Anal & Toxicol, BR-05508000 Sao Paulo - Brazil
 Univ Sao Paulo, Biochem & Pharmaceut Technol Dept, BR-05508000 Sao Paulo - Brazil
Total Affiliations: 4
Protein Expression and Purification;
Web of Science Citations:
In this work, the purification of a single-chain variable fragment (scFv) of an antibody by using liquid-liquid extraction in aqueous micellar two-phase systems was optimized by means of central composite design. Protein partitioning assays were performed by using the selected system composition in previous works: Triton X-114 at 4% wt/wt, yeast fermentation supernatant at 60% wt/wt, McIlvaine buffer pH 7.00. The other system component concentrations, Cibacron Blue F3GA (CB), Fabsorbent (TM) F1P HF (HF) and NaCl, were selected as independent variables. ScFv recovery percentage (%R) and purification factor (PF) were selected as the responses. According to the optimization process both, scFv recovery percentage and purification factor were favored with the addition of HF and NaCl in a range of concentrations around the central point of the second central composite design (HF 0.0120% w/w, CB 0.0200% w/w, NaCl 0.200% w/w). These experimental conditions allowed the concentration and pre-purification of scFv in the micelle-rich bottom phase of the systems with a recovery percentage superior to 88% and a purification factor of approximately 3.5. These results improved the previously presented works and demonstrated the convenience of using aqueous micellar two-phase systems as a first step in the purification of scFv molecules. (C) 2015 Elsevier Inc. All rights reserved. (AU)