title: Purification of equine whole IgG snake antivenom by using an aqueous two phase system as a primary purification step.
abstract: There is a need to introduce innovations in the manufacture of snake antivenoms to increase the supply of these products worldwide. In this work, the fractionation of equine hyperimmune plasma with a new methodology that includes an aqueous two phase system (ATPS) as a primary purification step was compared with the traditional method of caprylic acid precipitation. Hyperimmune plasma from horses immunized with the venoms of three snakes from sub-Saharan Africa was used as starting material for the production of both formulations. After being adjusted to the same lethal neutralizing activity, both antivenoms were compared in terms of their immunoreactivity, neutralization of in vitro venom activities, physicochemical characteristics, and stability. Their performance in terms of yield and purity was also assessed. The neutralization profile of in vitro enzymatic activities and the immunoreactivity, analyzed by ELISA and antivenomic approaches, were very similar for both preparations. Likewise, they behaved similarly in stability studies. However, ATPS-fractionated antivenom showed improved physicochemical profile and immunochemical purity and yield, mainly owing to its lower protein content. Additionally, this methodology allowed the recovery of albumin as a byproduct. ATPS purification constitutes a promising technology for antivenom production and should be further evaluated at preclinical and clinical levels.
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica. Electronic address: mariangela.vargasarroyo@ucr.ac.cr.
authors.forename: Mariángela
authors.initials: M
authors.lastname: Vargas
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Álvaro
authors.initials: Á
authors.lastname: Segura
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Mauren
authors.initials: M
authors.lastname: Villalta
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: María
authors.initials: M
authors.lastname: Herrera
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: José María
authors.initials: JM
authors.lastname: Gutiérrez
authors.affiliation: Instituto Clodomiro Picado, Facultad de Microbiología, Universidad de Costa Rica, San José, Costa Rica.
authors.forename: Guillermo
authors.initials: G
authors.lastname: León
mesh_terms: D000818:Animals; D000997:Antivenins / Q000276:immunology* / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006736:Horses; D007074:Immunoglobulin G / Q000276:immunology*; D014867:Water
chemical_list: D000997:Antivenins; D007074:Immunoglobulin G; D014867:Water
keywords: Aqueous two phase system; Caprylic acid; Immunoglobulins; Snake antivenom