Evaluation of a PEG/hydroxypropyl starch aqueous two‐phase system for the separation of monoclonal antibodies from cell culture supernatant

Abstract

In this study, an aqueous two-phase system (ATPS) with PEG and hydroxypropyl starch (HPS) was used to separate monoclonal antibody (mAb) from Chinese hamster ovary cell culture supernatant. The phase diagram of the PEG/HPS ATPS was determined, and the effects of NaCl addition were investigated. The results showed that NaCl addition could lead to a shift of the binodal curve and that phase separation would occur at higher PEG and HPS concentrations. The effects of NaCl addition, pH, and the load of cell supernatant on the partitioning of mAb in a PEG/HPS ATPS were investigated. It was found that with 6% cell supernatant and 15% NaCl addition at pH 6.0, the yield of mAb in the upper phase was 96.7% with a purity of 96.0%. The back-extraction of mAb with a PEG/phosphate ATPS were also studied, and the results showed that after the two-step extraction with ATPSs the purity of mAb could reach 97.6 ± 0.5% with a yield of 86.8 ± 1.0%, which was comparable to the purification with Protein A chromatography. These results indicate that the two-step extraction with PEG/HPS and PEG/phosphate ATPSs might be a promising alternative for the separation of mAb from cell culture supernatant.

Downstream processing of virus‐like particles with aqueous two‐phase systems: Applications and challenges

“…ATPSs have been long explored as a unique and efficient extraction pipeline, which offers a single-step procedure for the separation, recovery, purification, and enrichment of biomolecules such as enzymes, cells, particles [35], peptides, proteins [36][37][38][39][40][41], antibodies [42,43], pigments [44][45][46][47], algae-derived bioproducts [48], and natural products [49][50][51][52][53].…”

“…Aqueous two‐phase extraction (ATPE), which based on ATPSs, is a promising downstream processing technology in biotechnological industry and it has developed as new prospects in biomedical applications [13, 33, 34]. ATPSs have been long explored as a unique and efficient extraction pipeline, which offers a single‐step procedure for the separation, recovery, purification, and enrichment of biomolecules such as enzymes, cells, particles [35], peptides, proteins [36–41], antibodies [42, 43], pigments [44–47], algae‐derived bioproducts [48], and natural products [49–53].…”

Technology advancements in antibody purification

“…The authors found that the use of the ATPS protocol was highly useful for initial protein recovery and partial purification of mAb before proceeding to Protein-A affinity chromatography. Wu et al 103 reported the use of ATPS for the purification of mAbs in cell culture supernatant. The authors found that using the salt/ polymer method of ATPS (which involved increasing the concentration of NaCl) caused the formation of IgG precipitants, an undesirable end product contaminant.…”

“…101,103,104 Cao et al 104 investigated the purification of monoclonal anti-clenbuterol (CL) antibody using ATPS with differing PEG concentrations, pH values, and NaCl concentrations. Optimum conditions achieved antibody recovery yields of 87.86%.…”

The Research and Mechanism of Extracting Vitamin B6 Using Aqueous Two-Phase Systems

“…ey have mild separation conditions, rapid balance processes, and high rate of product recovery. As a "green chemistry," ATPS now is widely used in the separation of protein [6,7], DNA [8], drugs [9], antibiotic [10,11], and metal ion [12]. So, we pay more attention to find a new ATPS to extract vitamin B 6 .…”