title: New method to produce equine antirabies immunoglobulin F(ab')₂ fragments from crude plasma in high quality and yield.
abstract: Rabies is still a major cause of human deaths in several developing countries. According to the World Health Organization, administration of antirabies serum or antirabies immunoglobulin is recommended for patients who have experienced a category-III exposure to rabies. Improvement of antirabies immunoglobulin production is required to enhance safety and efficacy of the products. In this paper, a new method to produce equine antirabies immunoglobulin F(ab')(2) fragments from crude plasma is proposed. First, protein G affinity chromatography was used to purify IgG from equine plasma. Moreover, purification of IgG was shown to facilitate its digestion by pepsin. Compared to the direct digestion of crude plasma, a lower amount of pepsin and a shorter digestion time were required to completely digest the purified IgG to F(ab')(2). Complete digestion of purified IgG to F(ab')(2) was achieved at a pepsin/IgG (w/w) ratio of 5:45 with preservation of structure and potency. Finally, purification of F(ab')(2) was accomplished by a combination of protein A affinity chromatography and ultrafiltration with a 50-kDa nominal molecular weight cut-off membrane. The new process resulted in 68.9±0.6 (%) total recovery of F(ab')(2) and a F(ab')(2) product of high potency.
authors.affiliation: Department of Manufacturing Pharmacy, Faculty of Pharmacy, Mahidol University, Bangkok, Thailand.
authors.forename: Sukanda
authors.initials: S
authors.lastname: Kittipongwarakarn
authors.forename: Andrea
authors.initials: A
authors.lastname: Hawe
authors.forename: Ruedeeporn
authors.initials: R
authors.lastname: Tantipolphan
authors.forename: Kornvika
authors.initials: K
authors.lastname: Limsuwun
authors.forename: Sumana
authors.initials: S
authors.lastname: Khomvilai
authors.forename: Satit
authors.initials: S
authors.lastname: Puttipipatkhachorn
authors.forename: Wim
authors.initials: W
authors.lastname: Jiskoot
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D006736:Horses / Q000276:immunology*; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000097:blood / Q000302:isolation & purification; D011820:Rabies virus / Q000276:immunology*; D013056:Spectrophotometry, Ultraviolet; D014462:Ultrafiltration
chemical_list: D007140:Immunoglobulin Fab Fragments