A robust robotic high-throughput antibody purification platform.
Monoclonal antibodies (mAbs) have become the fastest growing segment in the drug market with annual sales of more than 40 billion US$ in 2013. The selection of lead candidate molecules involves the generation of large repertoires of antibodies from which to choose a final therapeutic candidate. Improvements in the ability to rapidly produce and purify many antibodies in sufficient quantities reduces the lead time for selection which ultimately impacts on the speed with which an antibody may transition through the research stage and into product development. Miniaturization and automation of chromatography using micro columns (RoboColumns(®) from Atoll GmbH) coupled to an automated liquid handling instrument (ALH; Freedom EVO(®) from Tecan) has been a successful approach to establish high throughput process development platforms. Recent advances in transient gene expression (TGE) using the high-titre Expi293F™ system have enabled recombinant mAb titres of greater than 500mg/L. These relatively high protein titres reduce the volume required to generate several milligrams of individual antibodies for initial biochemical and biological downstream assays, making TGE in the Expi293F™ system ideally suited to high throughput chromatography on an ALH. The present publication describes a novel platform for purifying Expi293F™-expressed recombinant mAbs directly from cell-free culture supernatant on a Perkin Elmer JANUS-VariSpan ALH equipped with a plate shuttle device. The purification platform allows automated 2-step purification (Protein A-desalting/size exclusion chromatography) of several hundred mAbs per week. The new robotic method can purify mAbs with high recovery (>90%) at sub-milligram level with yields of up to 2mg from 4mL of cell-free culture supernatant.
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia. Electronic address: Peter.Schmidt@csl.com.au.
Peter M
PM
Schmidt
Perkin Elmer, 530-540 Springvale Road, Glen Waverley, Victoria 3150, Australia.
Michael
M
Abdo
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Rebecca E
RE
Butcher
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Min-Yin
MY
Yap
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Pierre D
PD
Scotney
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Melanie L
ML
Ramunno
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Genevieve
G
Martin-Roussety
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Catherine
C
Owczarek
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Matthew P
MP
Hardy
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Chao-Guang
CG
Chen
CSL Limited, BIO21 Institute, 30 Flemington Road, Parkville, Victoria 3010, Australia.
Louis J
LJ
Fabri
D000911:Antibodies, Monoclonal / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D001331:Automation; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D057809:HEK293 Cells; D006801:Humans; D008904:Miniaturization; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D012371:Robotics; D013205:Staphylococcal Protein A / Q000737:chemistry / Q000378:metabolism
D000911:Antibodies, Monoclonal; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A
Antibody; High throughput; Liquid handling; Preparative purification; Protein A; Robotics