Abstract
The nematodeHaemonchus placeiis a pathogenic parasite, the most seriously affecting ruminant's health and being responsible for enormous economic losses all over the world. The present protocol describes different in vitro techniques to select potential candidate antigens with immune-protective activity from excretory and secretory products (ESP) fromH. placeitransitory infective larvae (xL3). ESP from xL3were obtained from the in vitro infective larvae (L3) maintained in Hank's medium at 37 °C with 5% CO2for 48 h. Then, the presence of ESP proteins was confirmed by SDS-PAGE to be used in an in vitro proliferation assay with bovine peripheral blood mononuclear cells (PBMCs). The ESP were exposed to the PBMCs during two different periods (24 and 48 h). Genes associated with immune response against the nematode were analyzed using relative gene expression and bioinformatic tools. These are simple, economic, and helpful tools to identify potential immune-protective molecules under in vitro conditions for confirming the efficacy of future in vivo assays. Graphical overview.