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Issue 17, 2014
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Affinity-based precipitation via a bivalent peptidic hapten for the purification of monoclonal antibodies

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Abstract

In a previous study, we demonstrated a non-chromatographic affinity-based precipitation method, using trivalent haptens, for the purification of mAbs. In this study, we significantly improved this process by using a simplified bivalent peptidic hapten (BPH) design, which enables facile and rapid purification of mAbs while overcoming the limitations of the previous trivalent design. The improved affinity-based precipitation method (ABPBPH) combines the simplicity of salt-induced precipitation with the selectivity of affinity chromatography for the purification of mAbs. The ABPBPH method involves 3 steps: (i) precipitation and separation of protein contaminants larger than immunoglobulins with ammonium sulfate; (ii) selective precipitation of the target-antibody via BPH by inducing antibody-complex formation; (iii) solubilization of the antibody pellet and removal of BPH with membrane filtration resulting in the pure antibody. The ABPBPH method was evaluated by purifying the pharmaceutical antibody trastuzumab from common contaminants including CHO cell conditioned media, DNA, ascites fluid, other antibodies, and denatured antibody with >85% yield and >97% purity. Importantly, the purified antibody demonstrated native binding activity to cell lines expressing the target protein, HER2. Combined, the ABPBPH method is a rapid and scalable process for the purification of antibodies with the potential to improve product quality while decreasing purification costs.

Graphical abstract: Affinity-based precipitation via a bivalent peptidic hapten for the purification of monoclonal antibodies

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Supplementary files

Article information


Submitted
01 May 2014
Accepted
12 Jun 2014
First published
12 Jun 2014

Analyst, 2014,139, 4247-4255
Article type
Paper
Author version available

Affinity-based precipitation via a bivalent peptidic hapten for the purification of monoclonal antibodies

M. W. Handlogten, J. F. Stefanick, P. E. Deak and B. Bilgicer, Analyst, 2014, 139, 4247
DOI: 10.1039/C4AN00780H

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