Purification of anti‐colorectal cancer monoclonal antibody CO17‐1A from insect cell culture using a French press and sonication |
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| Authors: | Chae‐Yeon Lim Se‐Ra Park Jeong‐Hwan Lee Kisung Ko |
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| Affiliation: | Department of Medicine, Medical Research Institute, College of Medicine, Chung‐Ang University, Seoul, Korea |
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| Abstract: | Monoclonal antibody (mAb) CO17‐1A binds to GA733, which is a tumor‐associated glycoprotein antigen highly expressed on the colorectal cancer cell surface. Thus, mAb CO17‐1A is considered a useful biomolecule for diagnosis and treatment against colorectal cancer. Previously, we established a baculovirus–insect cell expression system for the production of mAb CO17‐1A. In order to use mAb CO17‐1A as a diagnostic and therapeutic tool, however, the antibody must be properly purified from the insect cells. In this study, our aim was to investigate effective purification processes of mAb CO17‐1A expressed in Spodoptera frugiperda (Sf9) insect cells, using a French press and sonication for cell disruption. SDS‐PAGE confirmed that both mAb CO17‐1A and mAb CO17‐1A fused to the KDEL endoplasmic reticulum (ER) retention signal (mAb CO17‐1AK) were expressed clearly in Sf9 insect cells. Western blot analysis showed that detection levels of mAb CO17‐1A and CO17‐1AK were higher when the insect cells were disrupted two times by the French press and then sonicated, compared to only one French press disruption plus sonication. Optical microscopy confirmed that insect cells treated with both the French press and sonication were properly disrupted. Analysis of gene sequence information on mAb CO17‐1A verified that a signal peptide is present but a transmembrane protein does not exist. These results suggest that cell disruption by the French press twice and sonication once is an effective method for improving purification efficiency. |
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| Keywords: | baculovirus expression system colorectal cancer French press insect‐derived antibody mAb CO17‐1A |
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