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Updated: Jun 30, 2026

Production of Monoclonal Antibodies Targeting Aminopeptidase N in the Porcine Intestinal Mucosal Epithelium
Published on: May 18, 2021
Anion binding mediated precipitation of a peptibody
Atul Saluja1, Shon Crampton, Eva Kras
1Process & Product Development, 1201 Amgen Court West, MS AW2-D/3152, Seattle, Washington 98119-3105, USA.
Purpose:
Understand the underlying mechanism governing the salt-induced precipitation of a basic (pI = 8.8) protein, Peptibody A (PbA), in acidic solutions.
Methods:
The rate, extent, and reversibility of PbA precipitation was monitored over 4-weeks as a function of pH (3.7-5.0), salt concentration (0-400 mM), and ion identity using a series of monovalent, Hofmeister anions (F(-), Cl(-), Br(-), I(-), ClO(4) (-), SCN(-)) and cations (Li+, Na+, K+, Rb+, Cs+). The effects of salt on conformational stability and reduced valence were determined using Fourier-transform infrared spectroscopy, circular dichroism, and capillary electrophoresis/analytical ultracentrifugation.
Results:
PbA precipitation occurred upon salt addition and could be modulated with solution pH, salt identity & concentration. The precipitation was sensitive to anions, but not cations, and increased with anion size. A reverse Hofmeister effect (SCN(-) approximately ClO(4) (-)>I(-)>Cl(-)>Br(-)>F(-)) was observed with "salting-in" anions being the more effective precipitants. An increase in the precipitation rate below pH 4.3 indicated that protonation of aspartyl and glutamyl side-chains was also important for precipitation. The reversibility of precipitation was excellent (100%) at 4 degrees C but decreased upon storage at 25 degrees C and 37 degrees C; the loss in reversibility correlated with an increase in intermolecular beta-sheet content of the precipitate.
Conclusion:
Salts, employed as buffering, tonicifying, and viscosity modifying agents, may adversely affect the solubility of basic proteins formulated under acidic conditions.
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