Fig. 1

Production and selection of anti-OROV monoclonal antibodies. In A, scheme of inoculations in mice to produce mAbs, together with sera collection. In B, the screening of hybridomas resulting from the fusion is demonstrated, in addition to the following steps for selecting the most promising ones. In C, representative IFA for detection of anti-OROV antibodies from clones 63B3E7 (1:800) and 268B8E3 (1:100), together with the control sera (pre-immune and polyclonal, diluted at 1:100 and 1:800, respectively). The culture supernatant of the LD2 clones was purified and evaluated for the detection of viral antigens in C6/36 cells (1 × 10⁴ cells per well) uninfected (MOCK) and infected with OROV (MOI 1 for 48 h). In blue, cell nuclei stained with DAPI and in green OROV labeled with the respective mAb or polyclonal immune sera, followed by anti-mouse Ig conjugated with Alexa Fluor 488. In D–F, infected (OROV– red) and uninfected (MOCK - C6/36) cells– blue) were used for titration of polyclonal mouse sera (D), mAb 63B3E7 (E), and mAb 268B8A3 (F). The scale bar corresponds to 100 μm. D–F represents three experiments of three biological replicas