title: Development and validation of a quantitative competitive ELISA for potency testing of equine anti rabies sera with other potential use.
abstract: In case of a bite by a rabies infected animal, the World Health Organisation recommends a prophylactic treatment including the administration of Human Rabies Immunoglobulins (HRIGs) or highly purified F(ab')2 fragments produced from Equine Rabies Immunoglobulin (F(ab')2 - ERIGs). According to international regulation, quality control of F(ab')2 - ERIGs lots requires potency testing by the in vivo Mouse Neutralisation Test (MNT) prior marketing. However, the strategy of the 3Rs (Reduce, Refine, Replace) for animal testing required by the European Directive encourages the replacement of the in vivo potency test by an in vitro assay. In this context, a competitive ELISA method (c-ELISA) has been developed by the Agence Nationale de Sécurité du Médicament et des Produits de Santé where F(ab')2 - ERIGs are in competition with a monoclonal antibody recognizing the trimeric native form of the rabies glycoprotein. After a full validation study, the c-ELISA has been applied to commercial batches of F(ab')2 - ERIGs. A correlation study with the MNT demonstrated a similarity between the two methods (r=0.751). Moreover, the c-ELISA method which does not need any species specific reagent has been applied to HRIGs potency testing as an alternative method to Rapid Fluorescent Focus Inhibition Test (RFFIT), thus avoiding the handling of live rabies virus in BSL3 containment. In conclusion, the c-ELISA has shown its potential to replace MNT and possibly RFFIT for the quantification of rabies immunoglobulin. After optimisation it may be used for the quantification of rabies immunoglobulin in any animal species, notably for rabies immunogenicity assay in mice.
authors.affiliation: Agence Nationale de Sécurité du Médicament et des Produits de Santé Laboratory Controls Division, Batch Release and Marketing Surveillance of Biological Products Department, 321 Avenue Jean Jaurès, F-69007 Lyon, France.
authors.forename: Jehanara
authors.initials: J
authors.lastname: Korimbocus
authors.affiliation: Biological Controls of Immunological Products and Biological Security Department, 321 Avenue Jean Jaurès, F-69007 Lyon, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Dehay
authors.affiliation: Antiviral Strategies Unit, Virology Department, Institut Pasteur, 25, rue du Docteur Roux, F-75724 Paris Cedex 15, France.
authors.forename: Noël
authors.initials: N
authors.lastname: Tordo
authors.affiliation: Agence Nationale de Sécurité du Médicament et des Produits de Santé Laboratory Controls Division, Batch Release and Marketing Surveillance of Biological Products Department, 321 Avenue Jean Jaurès, F-69007 Lyon, France.
authors.forename: François
authors.initials: F
authors.lastname: Cano
authors.affiliation: Agence Nationale de Sécurité du Médicament et des Produits de Santé Laboratory Controls Division, Batch Release and Marketing Surveillance of Biological Products Department, 321 Avenue Jean Jaurès, F-69007 Lyon, France. Electronic address: sylvie.morgeaux@ansm.sante.fr.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Morgeaux
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D057134:Antibodies, Neutralizing / Q000276:immunology; D000914:Antibodies, Viral / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006736:Horses / Q000276:immunology; D007106:Immune Sera / Q000276:immunology*; D051379:Mice; D009500:Neutralization Tests; D011818:Rabies
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D007106:Immune Sera
keywords: 3Rs; Batch release; Competitive ELISA; Quality control; Rabies immunoglobulins (ERIGs, HRIGs); Validation