Automated 2D-HPLC method for characterization of protein aggregation with in-line fraction collection device.
Monoclonal antibodies are mainly produced by mammalian cell culture, which due to its complexity, results in a wide range of product variants/isoforms. With the growing implementation of Quality by Design (QbD) and Process Analytical Technology (PAT) in drug manufacturing, monitoring and controlling quality attributes within a predefined range during manufacturing may provide added consistency to product quality. To implement these concepts, more robust analytical tools could reduce the time needed for monitoring quality attributes during upstream processing. The formation of protein aggregates is one such quality attribute that can lead to safety and efficacy issues in the final drug product. Described in this study is a fully automated two-dimensional high performance liquid chromatography (2D-HPLC) method for characterizing protein aggregation of crude in-process bioreactor samples. It combines protein A purification and separation by size exclusion into a single analytical module that has the potential to be employed at-line within a bioprocessing system. This method utilizes a novel in-line fraction collection device allowing for the collection of up to twelve fractions from a single sample or peak which facilitates the subsequent linked analysis of multiple protein peaks of interest in one chromatography module.
Food and Drug Administration, Center for Drug Evaluation and Research, Office of Product Quality, Office of Biotechnology Products, Division of Biotechnology Review and Research II, Silver Spring, MD, United States.
Abasha
A
Williams
Food and Drug Administration, Center for Drug Evaluation and Research, Office of Product Quality, Office of Biotechnology Products, Division of Biotechnology Review and Research II, Silver Spring, MD, United States.
Erik K
EK
Read
Food and Drug Administration, Center for Drug Evaluation and Research, Office of Product Quality, Office of Biotechnology Products, Division of Biotechnology Review and Research II, Silver Spring, MD, United States.
Cyrus D
CD
Agarabi
Food and Drug Administration, Center for Drug Evaluation and Research, Office of Product Quality, Office of Biotechnology Products, Division of Biotechnology Review and Research II, Silver Spring, MD, United States.
Scott
S
Lute
Food and Drug Administration, Center for Drug Evaluation and Research, Office of Product Quality, Office of Biotechnology Products, Division of Biotechnology Review and Research II, Silver Spring, MD, United States. Electronic address: kurt.brorson@fda.hhs.gov.
Kurt A
KA
Brorson
D000911:Antibodies, Monoclonal / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification; D002850:Chromatography, Gel / Q000295:instrumentation / Q000379:methods; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation* / Q000379:methods*; D004867:Equipment Design; D066329:Protein Aggregates; D013205:Staphylococcal Protein A / Q000032:analysis / Q000737:chemistry / Q000302:isolation & purification
D000911:Antibodies, Monoclonal; D066329:Protein Aggregates; D013205:Staphylococcal Protein A
2D-HPLC; Heart-cutting; Monoclonal antibody purification; PAT; Protein aggregation; QBD