• Standardization and Statistical validation under ISO/IEC 17025 standards of the INCUINTA ELISA to detect antibodies against BoHV-1 in bovine and guinea pig serum

PARREÑO, VIVIANA; Y COL.

Buenos Aires

Simposio; Symposium: Practical Alternatives to reduce animal testing in quality control of veterinary biologicals in the Americas; 2010

IABS-OIE

Standardization and Statistical Validation under ISO/IEC 17025 standards of the INCUINTA ELISA to detect antibodies against BoHV-1 in Bovine and Guinea Pig serum. Parreño, Viviana1,2* Romera, Alejandra,1,2* ; Makek, Lucia3; Rodriguez, Daniela1 ; Malacari, Dario1;  Maidana Silvina1; Compaired Diego1; Combessies, Gustavo4; Vena, Maria Marta5 ; Garaicoechea, Lorena1; Wigdorovitz, Andrés1; Marangunich, Laura3 ; and Fernandez, Fernando1. * Both authors contributed equally to this work.   vparreno@cnia.inta.gov.ar 1 Instituto de Virología, CICV y A – INTA. 2 CONICET. 3  Universidad Nacional de Tres de Febrero, UNTREF. 4 Laboratorio Azul. 5 BIOGENESIS-BAGO S.A., Argentina. Two indirect ELISAs to quantify antibodies (Ab) to BoHV-1 in serum of infected or vaccinated cattle and immunized guinea pig used for vaccine potency testing were developed and validated. The cut off of the assays was established at 40 PP%, for both species. The reliability of the assays, given by their diagnosis sensitivity (DSe) and specificity (DSp) was 89.7 and 100% for bovine and 95 and 100% for guinea pigs, respectively. The relative Se and Sp for guinea pigs, regarding the virus neutralization assay (VN), was RSe=94.95% and RSp= 99.21%. There was a good correlation between the VN and ELISA Ab titers from the serum of the same bovine (R=0.85; p<0.0001) and the same guinea pig (R=0.92; p<0.0001). A similar correlation was observed when analyzing the mean Ab titers of groups of vaccinated animals (for bovine, R=0.84 and for guinea pigs, R=0.91; p<0.0001). Inter-laboratory assays including 4 laboratories for bovine and 3 laboratories for guinea pigs samples, gave very good reproducibility (CV=12.4% for bovine and CV=→0 for guinea pigs),  fulfilling the requirements of the OIE (CV<30%). The validated ELISAs will contribute to control BoHV-1 infection in Argentina and will be used for selection of BoHV-1 free bovines for international trade and vaccine potency testing in the host species and in a lab animal model.