Affinity chromatography as a tool for antibody purification

• Published in: Methods (San Diego, Calif.) Calif.), 2012-02, Vol.56 (2), p.116-129
• Main Authors: Ayyar, B. Vijayalakshmi, Arora, Sushrut, Murphy, Caroline, O’Kennedy, Richard
• Format: Article
• Language: English
• Subjects: Affinity chromatography; Affinity ligands; Affinity matrices; Animals; Antibodies - chemistry; Antibodies - classification; Antibodies - isolation & purification; Antibody; Antibody Affinity; Antibody purification; Antibody Specificity; Antigens - chemistry; Chromatography, Affinity - methods; Chromatography, Affinity - standards; Culture Media - chemistry; Humans; Ligands; Reproducibility of Results; Sensitivity and Specificity; Time Factors
• ISSN: 1046-2023; 1095-9130
• DOI: 10.1016/j.ymeth.2011.10.007

The global antibody market has grown exponentially due to increasing applications in research, diagnostics and therapy. Antibodies are present in complex matrices (e.g. serum, milk, egg yolk, fermentation broth or plant-derived extracts). This has led to the need for development of novel platforms for purification of large quantities of antibody with defined clinical and performance requirements. However, the choice of method is strictly limited by the manufacturing cost and the quality of the end product required. Affinity chromatography is one of the most extensively used methods for antibody purification, due to its high selectivity and rapidity. Its effectiveness is largely based on the binding characteristics of the required antibody and the ligand used for antibody capture. The approaches used for antibody purification are critically examined with the aim of providing the reader with the principles and practical insights required to understand the intricacies of the procedures. Affinity support matrices and ligands for affinity chromatography are discussed, including their relevant underlying principles of use, their potential value and their performance in purifying different types of antibodies, along with a list of commercially available alternatives. Furthermore, the principal factors influencing purification procedures at various stages are highlighted. Practical considerations for development and/or optimizations of efficient antibody-purification protocols are suggested.