Id screen bluetongue competition

All serum samples of field and experimental specimens were screened for group specific antibodies using a commercial cELISA targeting the VP7 (ID Screen^® Bluetongue Competition, ID-Vet, France) according to the manufacturer’s instructions [32 (link)]. Samples with ≤50% of negativity compared to the negative control (S/N) were considered as positive, samples with ≥50% S/N as negative.

All sera were tested for the presence of BTV-specific antibodies by means of a commercially available competitive ELISA (c-ELISA) (ID Screen® Blue Tongue Competition, ID VET, Montpellier, France) performed according to the instructions provided by the manufacturer. Results were expressed as a percentage negativity (PN) compared to the negative kit control and were classified into positive (PN ≤ 65), doubtful (PN > 65 but ≤ 75), and negative (PN > 75) results based on the optimal cut-off point for diagnostic purposes of 65 PN determined by Vandenbussche et al. (31 (link)).

Sera were separated from blood samples by centrifugation at 1000 g for 10 min and kept at -20 °C until testing. A commercial assay (ID Screen® Bluetongue Competition, ID Vet) was used to detect BTV antibodies in each sample. This competitive assay detects antibodies against all BTV serotypes (using the VP7 core protein as antigen) and is selective (does not cross-react with antibodies against the closely related epizootic hemorrhagic disease virus). The assay was performed following the manufacturer's instructions and optical density values at 450 nm were determined using a spectrophotometer. Results were calculated for each serum as a ratio to the mean of negative controls (Sample/Negative control ratios, or S/N) and compared to cut-off values (S/N < 40% are defined as positives).

At the IVI in Switzerland, the commercially available ELISA INgezim BTV DR 12.BTV.K.0 (INGENASA, Eurofins, Budapest, Hungary) targeting the VP7 was used according to the manufacturer’s protocol for analysis of the field plasma samples. Samples were considered positive when the OD value at 450 nm was higher than the cut-off (15% of positive control). Samples were considered negative if the OD value was equal or lower than the positive cut-off.
At the FLI in Germany, all serum samples from the animal experiments were screened for BTV-group-specific antibodies using a cELISA targeting the VP7 (ID Screen^® Bluetongue Competition, ID-Vet, France) according to the manufacturer’s instructions. The cut-off of 50% according to the manufacture’s instruction was applied. Samples with S/N ≤ 50% were considered positive, samples with >50% S/N as negative.