title: Development of an automated mid-scale parallel protein purification system for antibody purification and affinity chromatography.
abstract: Protein purification is often a bottleneck during protein generation for large molecule drug discovery. Therapeutic antibody campaigns typically require the purification of hundreds of monoclonal antibodies (mAbs) during the hybridoma process and lead optimization. With the increase in high-throughput cloning, faster DNA sequencing, and the use of parallel protein expression systems, a need for high-throughput purification approaches has evolved, particularly in the midsize range between 20 ml and 100 ml. To address this we modified a four channel Gilson solid phase extraction system (referred to as MG-SPE) with switching valves and sample holding loops to be able to perform standard affinity purification using commercially available columns and micro-titer format deep well blocks. By running 4 samples in parallel, the MG-SPE has the capacity to purify up to 24 samples of greater than 50 ml each using a single-step affinity purification protocol or a two-step protocol consisting of affinity chromatography followed by desalting/buffer exchange overnight (∼12 h run time). Our evaluation of affinity purification using mAbs and Fc-fusion proteins from mammalian cell supernatants demonstrates that the MG-SPE compared favorably with industry standard systems for both protein quality and yield. Overall the system is simple to operate and fills a void in purification processes where a simple, efficient, automated system is needed for affinity purification of midsize research samples.
authors: Department of Chemistry, University of Missouri-Kansas City, Kansas City, MO 64110, USA.
authors: Chi
authors: C
authors: Zhang
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Alexander M
authors: AM
authors: Long
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Brooke
authors: B
authors: Swalm
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Ken
authors: K
authors: Charest
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Yan
authors: Y
authors: Wang
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Jiali
authors: J
authors: Hu
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Craig
authors: C
authors: Schulz
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA.
authors: Wolfgang
authors: W
authors: Goetzinger
authors: Therapeutic Discovery, Amgen Inc., 360 Binney St., Cambridge, MA 02141, USA. Electronic address: brianhall22@gmail.com.
authors: Brian E
authors: BE
authors: Hall
mesh_terms: D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods*; D057809:HEK293 Cells; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000302:isolation & purification*
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G
keywords: Affinity chromatography; Automation; High-throughput protein purification; Monoclonal antibody (mAb)