Antibody capture by mixed-mode chromatography: A comprehensive study from determination of optimal purification conditions to identification of contaminating host cell proteins

Purification process of recombinant monoclonal antibodies with mixed mode chromatography

“…Identification was carried out by database search against the Uniprot entries from Cricetulus griseus. To date, several mixed mode resins have been tested for antibody capture [23,24]. Both PPA and HEA HyperCel resins gave good results in these studies so we examined the following characteristics to choose the most appropriate one: yield, elimination of HCPs and pH for elution.…”

“…Conditions for binding and elution were tested in microplates as described previously [23,28]. Optimal conditions were found to be 50 mM sodium citrate buffer pH 4 (6 mS/cm) for equilibration and 200 mM sodium phosphate buffer at pH 7.5 and 21 mS/cm for elution.…”

“…Optimal conditions for mAb capture by HEA HyperCel were previously defined [23]. Briefly, 0.1 M sodium phosphate buffer pH 7.3 and 13 mS/cm were used for equilibration and binding, then 0.1 M sodium phosphate pH 6.5 and 15 mS/cm were used as intermediate buffer (wash step) to remove any weakly adsorbed contaminants.…”

Media Selection in Ion-Exchange Chromatography in a Single Microplate

“…We have done screening of chromatographic conditions in microplate with a design of experiment approach to purify antibody from CHO cell culture. We tested both different resins and different conditions for wash and elution steps [ 8 ]. Unfortunately resin manufacturers do not provide microplates with different resins made by them and by competitors.…”

Protein Adsorption and Transport in Novel Chromatographic Anion Exchange and Multimodal Resins

“…As a result, developing processes based on MMC usually requires extensive experimentation [14,35]. A better understanding of both protein-ligand interactions and of the transport mechanism in MMC is thus desirable in order to develop more rational approaches to process development.…”