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Evaluation of the Efficiency of Protein A Affinity Chromatography to Purify a Monoclonal Antibody for Cancer Treatment and its Purity Analysis

[ Vol. 7 , Issue. 2 ]


Ayesha Akhtar*, Shivakumar Arumugam and Shoaib Alam   Pages 121 - 133 ( 13 )


Background: Protein A affinity chromatography is often employed as the most crucial purification step for monoclonal antibodies to achieve high yield with purity and throughput requirements.

Introduction: Protein A, also known as Staphylococcal protein A (SPA) is found in the cell wall of the bacteria staphylococcus aureus. It is one of the first discovered immunoglobulin binding molecules and has been extensively studied since the past few decades. The efficiency of Protein A affinity chromatography to purify a recombinant monoclonal antibody in a cell culture sample has been evaluated, which removes 99.0% of feed stream impurities.

Materials and Method: We have systematically evaluated the purification performance by using a battery of analytical methods SDS-PAGE (non-reduced and reduced sample), Cation Exchange Chromatography (CEX), Size-exclusion chromatography (SEC), and Reversed phased-Reduced Chromatography for a CHO-derived monoclonal antibody.

Results and Discussion: The analytical test was conducted to determine the impurity parameter, Host Cell Contaminating Proteins (HCP). It was evaluated to be 0.015ng/ml after the purification step; while initially, it was found to be 24.431ng/ml.

Conclusion: The tests showed a distinct decrease in the level of different impurities after the chromatography step. It can be concluded that Protein A chromatography is an efficient step in the purification of monoclonal antibodies.


Affinity chromatography, monoclonal antibody, protein A, SDS-PAGE, high-performance liquid chromatography (HPLC), host cell contaminating proteins (HCP).


Department of Biotechnology, VIT University, Vellore, School of Bioscience and Technology, VIT University, Vellore, Research and Development, Jamjoom Pharmaceuticals, Jeddah

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