Purification of anti-Japanese encephalitis virus monoclonal antibody by ceramic hydroxyapatite chromatography without proteins A and G.
Antibody purification using proteins A and G has been a standard method for research and industrial processes. The conventional method, however, includes a three-step process, including buffer exchange, before chromatography. In addition, proteins A and G require low pH elution, which causes antibody aggregation and inactivates the antibody's immunity. This report proposes a two-step method using hydroxyapatite chromatography and membrane filtration, without proteins A and G. This novel method shortens the running time to one-third the conventional method for each cycle. Using our two-step method, 90.2% of the monoclonal antibodies purified were recovered in the elution fraction, the purity achieved was >90%, and most of the antigen-specific activity was retained. This report suggests that the two-step method using hydroxyapatite chromatography and membrane filtration should be considered as an alternative to purification using proteins A and G.
R&D Department, Pentax New Ceramics Division, Hoya Corporation, 1-1-110 Tsutsujigaoka,Akishima-shi, Tokyo, Japan. maiko.saito@pc.hoya.co.jp
Maiko
M
Saito
Yae
Y
Kurosawa
Tsuneo
T
Okuyama
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000302:isolation & purification*; D000914:Antibodies, Viral / Q000276:immunology / Q000302:isolation & purification*; D002852:Chromatography, Ion Exchange / Q000379:methods*; D017886:Durapatite / Q000737:chemistry; D004664:Encephalitis Virus, Japanese / Q000276:immunology*; D051379:Mice; D008851:Micropore Filters
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D017886:Durapatite