Capillary electrophoresis-based assessment of nanobody affinity and purity

R. Haselberg, S. Oliveira, R. van der Meel, G.W. Somsen, G.J. de Jong

Research output: Contribution to JournalArticleAcademicpeer-review

Abstract

Drug purity and affinity are essential attributes during development and production of therapeutic proteins. In this work, capillary electrophoresis (CE) was used to determine both the affinity and composition of the biotechnologically produced "nanobody" EGa1, the binding fragment of a heavy-chain-only antibody. EGa1 is an antagonist of the epidermal growth factor receptor (EGFR), which is overexpressed on the surface of tumor cells. Using a background electrolyte (BGE) of 50mM sodium phosphate (pH 8.0) in combination with a polybrene-poly(vinylsulfonic acid) capillary coating, CE analysis of EGa1 showed the presence of at least three components. Affinity of the EGa1 components towards the extracellular domain of EGFR was assessed by adding different concentrations (0-12nM) of the receptor to the BGE while measuring the effective electrophoretic mobility of the respective EGa1 components. Binding curves obtained by plotting electrophoretic mobility shifts as a function of receptor concentration, yielded dissociation constants (K
Original languageEnglish
Pages (from-to)1-6
JournalAnalytica Chimica Acta
Volume818
DOIs
Publication statusPublished - 2014

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