Optimized extraction of a single-chain variable fragment of antibody by using aqueous micellar two-phase systems

Autor(es)

Malpiedi, Luciana P. ^[1] ; Nerli, Bibiana B. ^[1] ; Taqueda, Maria E. S. ^[2] ; Abdalla, Dulcineia S. P. ^[3] ; Pessoa, Jr., Adalberto ^[4]

Afiliação do(s) autor(es)

^[1] Univ Nacl Rosario, Inst Proc Biotecnol & Quim IPROBYQ, Dept Quim Fis, RA-2000 Rosario, Santa Fe - Argentina

^[2] Univ Sao Paulo, Polytech Sch, Dept Chem Engn, BR-05508000 Sao Paulo - Brazil

^[3] Univ Sao Paulo, Dept Clin Anal & Toxicol, BR-05508000 Sao Paulo - Brazil

^[4] Univ Sao Paulo, Biochem & Pharmaceut Technol Dept, BR-05508000 Sao Paulo - Brazil

• Tipo de documento: Artigo Científico
• Fonte: Protein Expression and Purification; v. 111, p. 53-60, JUL 2015.
• Citações Web of Science: 3

Resumo

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)

• Processo FAPESP: 11/20521-0 - Extração de fragmentos de anticorpos monoclonais scFv por sistemas micelares de duas fases aquosas
• Beneficiário: Luciana Pellegrini Malpiedi
• Linha de fomento: Bolsas no Brasil - Pós-Doutorado