title: CZE for glycoform profiling and quality assessment of recombinant human interleukin-7.
abstract: The aim of the present work was to develop a simple high-resolution CZE method for glycoform profiling and quality assessment of recombinant human interleukin-7 (rhIL-7) produced in Chinese hamster ovary cells. Classical and isoelectric buffers and several monoamines used as alkaline component of BGE at very low concentration have been investigated in order to optimize the resolution. The best separation was achieved using a fused-silica capillary and a buffer composed of 25 mM citrate/triethanolamine at pH 2.6. Under these acidic conditions, triethanolamine was able to reverse EOF favorably and to limit rhIL-7 adsorption. This method allowed the separation of four groups of peaks ranging from low to high-sialylated glycoforms. An extensive study on inter-run rinsing procedures has been conducted. Rinsing with 50 mM SDS was retained to achieve the optimal repeatability. Excellent intermediate precision was obtained for migration time (RSD < 0.6%), while RSD for intraday studies were only less than 2.9%. Satisfactory inter and intraday repeatabilities were also observed for relative peak area. We finally demonstrated that reliable information could be obtained to address comparability studies and demonstrate batch-to-batch consistency of biomanufactured rhIL-7.
authors.affiliation: Univ Paris-Sud, Laboratoire de Protéines et Nanotechnologies en Sciences Séparatives, Faculté de Pharmacie, Châtenay-Malabry, France.
authors.forename: Youssef
authors.initials: Y
authors.lastname: Alahmad
authors.forename: Nguyet
authors.initials: N
authors.lastname: Thuy Tran
authors.forename: Jérémy
authors.initials: J
authors.lastname: Duboeuf
authors.forename: Anne
authors.initials: A
authors.lastname: Grégoire
authors.forename: Iann
authors.initials: I
authors.lastname: Rancé
authors.forename: Myriam
authors.initials: M
authors.lastname: Taverna
mesh_terms: D000818:Animals; D002021:Buffers; D016466:CHO Cells; D019343:Citric Acid / Q000737:chemistry; D006224:Cricetinae; D003412:Cricetulus; D019075:Electrophoresis, Capillary / Q000379:methods*; D004983:Ethanolamines / Q000737:chemistry; D006023:Glycoproteins / Q000737:chemistry*; D006031:Glycosylation; D006863:Hydrogen-Ion Concentration; D015851:Interleukin-7 / Q000737:chemistry*; D020033:Protein Isoforms; D011994:Recombinant Proteins / Q000737:chemistry; D015203:Reproducibility of Results
chemical_list: D002021:Buffers; D004983:Ethanolamines; D006023:Glycoproteins; C508605:IL7 protein, human; D015851:Interleukin-7; D020033:Protein Isoforms; D011994:Recombinant Proteins; D019343:Citric Acid; C009546:triethanolamine