[Validation of a real time RT-PCR assay to detect foot-and-mouth disease virus and assessment of its performance in acute infection].

A specific real time reverse transcription polymerase chain reaction (RT-PCRrt) for the detection of foot-and-mouth disease virus was validated using the LightCycler thermocycler 2.0 and its reagents as recommended by the World Organization for Animal Health and was assessed for the detection of the virus in acute infection of cattle experimentally vaccinated and challenged with virus A Argentina/2001 or A24 Cruzeiro. The technique proved to be robust, showing coefficients of variation lower than 4% for different ARN extractions, days or repetitions and was able to detect up to 0,4 TCID 50%, and/or up to 100 RNA molecules.

Validation of an indirect ELISA to detect antibodies against BoHV-1 in bovine and guinea-pig serum samples using ISO/IEC 17025 standards.

Two ELISAs to quantify antibodies to BoHV-1 in the sera of cattle and immunized guinea pigs were developed and validated using ISO/IEC 17025 standards. The cut-off value of the assay was established at 20% positivity of a high positive control for screening of cattle. Using this threshold, the assay properly classified the OIE bovine reference sera EU1, EU2 and EU3.

Novel purification method for recombinant 3AB1 nonstructural protein of foot-and-mouth disease virus for use in differentiation between infected and vaccinated animals.

An indirect enzyme-linked immunosorbent assay (ELISA) was developed for differentiation of animals infected with foot-and-mouth disease virus (FMDV) from vaccinated animals. The test was based on a highly pure and concentrated preparation of recombinant 3AB1 protein obtained by expression in a prokaryotic system, protein separation by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and electro elution. Experimental- and field-serum samples from naive, vaccinated, and infected cattle were tested for anti3AB1 antibody using the ELISA.