Purification of plant-derived anti-virus mAb through optimized pH conditions for coupling between protein A and epoxy-activated beadsopen access
- Authors
- Song, Ilchan; Kang, Yang Joo; Choi, Su-Lim; Han, Dalmuri; Kim, Deuk-Su; Lee, Hae Kyung; Lee, Joon-Chul; Park, Jeanho; Kim, Do-Sun; Ko, Kisung
- Issue Date
- May-2019
- Publisher
- PEERJ INC
- Keywords
- Antibody; Epoxy-activated beads; pH condition; Protein A; Purification
- Citation
- PEERJ, v.7, no.5
- Journal Title
- PEERJ
- Volume
- 7
- Number
- 5
- URI
- https://scholarworks.bwise.kr/cau/handle/2019.sw.cau/32770
- DOI
- 10.7717/peerj.6828
- ISSN
- 2167-8359
- Abstract
- The main goal of this research was to determine optimum pH conditions for coupling between protein A and epoxy-activated Sepharose beads for purification of monoclonal antibodies (mAbs) expressed in plants. To confirm the effect of pH conditions on purification efficacy, epoxy-activated agarose beads were coupled to protein A under the pH conditions of 8.5, 9.5, 10.5, and 11.5 (8.5R, 9.5R, 10.5R, and 11.5R, respectively). A total of 300 g of fresh leaf tissue of transgenic Arabidopsis expressing human anti-rabies mAb (mAb(P)) SO57 were harvested to isolate the total soluble protein (TSP). An equal amount of TSP solution was applied to five resin groups including commercial protein A resin (GR) as a positive control. The modified 8.5R, 9.5R, 10.5R, and 11.5R showed delayed elution timing compared to the GR control resin. Nano-drop analysis showed that the total amount of purified mAb(P) SO57 mAbs from 60 g of fresh leaf mass were not significantly different among 8.5R (400 mu g), 9.5R (360 mu g), 10.5R (380 mu g), and GR (350 mu g). The 11.5R (25 mu g) had the least mAb(P )SO57. SDS-PAGE analysis showed that the purity of mAb(P) SO57 was not significantly different among the five groups. Rapid fluorescent focus inhibition tests revealed that virus-neutralizing efficacies of purified mAb(P) SO57 from all the five different resins including the positive control resin were similar. Taken together, both pH 8.5 and 10.5 coupling conditions with high recovery rate should be optimized for purification of mAb(P )SO57 from transgenic Arabidopsis plant, which will eventually reduce down-stream cost required for mAb production using the plant system.
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