Direct isolation of monoclonal antibodies from tissue culture supernatant using the cation-exchange cellulose Express-Ion S.
The chromatography of the murine hybridoma cell C595/102 culture supernatant expressing the therapeutic monoclonal antibody C595, on the cation-exchange cellulose Whatman Express-Ion Exchanger S has been investigated. Initial method scouting studies using purified C595 in 1-ml mini columns demonstrated that binding capacity and binding efficiency were dependent not only on decreasing pH but also on the buffer salts used to prepare the mobile phase. Under optimised conditions of 0.1 M sodium acetate buffer, pH 5.0, we were able to separate purified C595 from BSA, the major contaminant in tissue culture fluid. Under these conditions immunoreactive C595 could be isolated directly from tissue culture supernatant. A scale-down study was carried out using a 25-ml column operated at a flow-rate of 150 cm/h which also yielded purified immunoreactive antibody. This procedure should now be suitable for scale-up.
Denton
G
G
Cancer Research Laboratories, School of Pharmaceutical Sciences, University of Nottingham, University Park, UK.
Murray
A
A
Price
M R
MR
Levison
P R
PR
D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D015153:Blotting, Western; D002411:Cation Exchange Resins; D002852:Chromatography, Ion Exchange / Q000295:instrumentation*; D046508:Culture Techniques; D004591:Electrophoresis, Polyacrylamide Gel; D006825:Hybridomas / Q000276:immunology; D051379:Mice
D000911:Antibodies, Monoclonal; D002411:Cation Exchange Resins