title: Continuous precipitation of IgG from CHO cell culture supernatant in a tubular reactor.
abstract: We successfully transferred a two-stage batch precipitation-based antibody capture step to continuous mode using continuous tubular reactors. The precipitation process solely employs a cheap mineral salt (CaCl2 ) and an organic solvent (ethanol) and could replace the costly protein A capture step in the purification of recombinant antibodies from cell culture supernatant. The time from startup untill attaining steady state conditions was reached in less than 15 minutes and both reactors were operated for several hours at steady state without manual intervention, delivering antibody at a constant yield and purity. An overall yield of > 90 percent, with a host cell protein reduction from 42 777 to 9000 ppm and a DNA reduction from 359 ppm to 7 ppm, could be achieved for the antibody investigated. The precipitated antibody can be dissolved at very high concentrations (> 40 g/L) in numerous buffer systems of various pH and high and low ionic strength, thereby rendering a subsequent concentration or buffer exchange step redundant. This system enables cell culture supernatants with low or high antibody titer to be processed with constant reactor size and without changing any parameters or increasing precipitant consumption. Aggregate levels were below 1% under all conditions tested. Purification by precipitation did not affect binding to CD16a or the isoform distribution of the antibody.
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.|Austria Centre for Industrial Biotechnology, Vienna, Austria.
authors.forename: Nikolaus
authors.initials: N
authors.lastname: Hammerschmidt
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
authors.forename: Beate
authors.initials: B
authors.lastname: Hintersteiner
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
authors.forename: Nico
authors.initials: N
authors.lastname: Lingg
authors.affiliation: Department of Biotechnology, University of Natural Resources and Life Sciences Vienna, Vienna, Austria. alois.jungbauer@boku.ac.at.|Austria Centre for Industrial Biotechnology, Vienna, Austria. alois.jungbauer@boku.ac.at.
authors.forename: Alois
authors.initials: A
authors.lastname: Jungbauer
mesh_terms: D000818:Animals; D019149:Bioreactors; D001709:Biotechnology / Q000379:methods*; D016466:CHO Cells; D002122:Calcium Chloride / Q000737:chemistry; D011232:Chemical Precipitation; D006224:Cricetinae; D003412:Cricetulus; D000431:Ethanol; D007074:Immunoglobulin G / Q000737:chemistry / Q000302:isolation & purification*; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification*; D013205:Staphylococcal Protein A
chemical_list: D007074:Immunoglobulin G; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A; D000431:Ethanol; D002122:Calcium Chloride
keywords: Antibody manufacturing; Capture; Downstream processing; Precipitation