Monoclonal antibody production using a new supermacroporous cryogel bioreactor.
A supermacroporous cryogel bioreactor has been developed to culture hybridoma cells for long-term continuous production of monoclonal antibodies (mAb). Hybridoma clone M2139, secreting antibodies against J1 epitope (GERGAAGIAGPK; amino acids, 551-564) of collagen type II, are immobilized in the porous bed matrix of a cryogel column (10 mL bed volume). The cells got attached to the matrix within 48 h after inoculation and grew as a confluent sheet inside the cryogel matrix. Cells were in the lag phase for 15 days and secreted mAb into the circulation medium. Glucose consumption and lactic acid production were also monitored, and during the exponential phase (approximately 20 days), the hybridoma cell line consumed 0.75 mM day-1 glucose, produced 2.48 mM day-1 lactic acid, and produced 6.5 microg mL-1 day-1 mAb during the exponential phase. The mAb concentration reached 130 microg mL-1 after continuous run of the cryogel column for 36 days. The yield of the mAb after purification was 67.5 mg L-1, which was three times greater than the mAb yield obtained from T-flask batch cultivation. Even after the exchange of medium reservoir, cells in the cryogel column were still active and had relatively stable mAb production for an extended period of time. The bioreactor was operated continuously for 55 days without any contamination. The results from ELISA as well as arthritis experiments demonstrate that the antibodies secreted by cells grown on the cryogel column did not differ from antibodies purified from the cells grown in commercial CL-1000 culture flasks. Thus, supermacroporous cryogels can be useful as a supporting material for productive hybridoma cell culture. Cells were found to be viable inside the porous matrix of the cryogel during the study period and secreted antibodies continuously. The antibodies thus obtained from the cryogel reactor were found to be functionally active in vivo, as demonstrated by their capacity to induce arthritis in mice.
Nilsang
Suthasinee
S
Department of Food Engineering and Bioprocess Technology, School of Environmental, Resources, and Development, Asian Institute of Technology, Klong Luang Pathumthani 12120, Thailand.
Nandakumar
Kutty Selva
KS
Galaev
Igor Yu
IY
Rakshit
Sudip Kumar
SK
Holmdahl
Rikard
R
Mattiasson
Bo
B
Kumar
Ashok
A
D000596:Amino Acids / Q000737:chemistry; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D001168:Arthritis / Q000378:metabolism; D019149:Bioreactors; D001709:Biotechnology / Q000295:instrumentation* / Q000379:methods; D001798:Blood Proteins / Q000737:chemistry*; D045744:Cell Line, Tumor; D002470:Cell Survival; D059985:Cryogels; D000939:Epitopes / Q000737:chemistry; D005353:Fibronectins / Q000737:chemistry*; D005780:Gelatin / Q000737:chemistry; D005947:Glucose / Q000737:chemistry; D006825:Hybridomas / Q000737:chemistry / Q000378:metabolism; D020100:Hydrogels; D007773:Lactates / Q000737:chemistry; D051379:Mice
D000596:Amino Acids; D000911:Antibodies, Monoclonal; D001798:Blood Proteins; D059985:Cryogels; D000939:Epitopes; D005353:Fibronectins; D020100:Hydrogels; D007773:Lactates; D005780:Gelatin; D005947:Glucose