Goat anti horse igg h l

Both rabbit and horse polyclonal antibodies raised against the venom of N. atra were donated by the Centers for Disease Control, Taiwan. These antivenoms were further affinity-purified by LTK BioLaboratories Co., Ltd. with an antigen-immobilized column system and dialyzed with PBS. Peroxidase-conjugated goat anti-rabbit IgG (H + L) and goat anti-horse IgG (H + L) were purchased from Jackson ImmunoResearch Inc. (West Grove, PA, USA). SureBlue Reserve™ TMB Microwell Peroxidase Substrate was purchased from SeraCare Life Sciences Inc. (Milford, MA, USA). Bolt™ 4–12% Bis-Tris Plus Gels and Novex^® Sharp Pre-stained Protein Standard were purchased from Thermo Fisher Scientific Inc. (Waltham, MA, USA). Immobilon-PSQ PVDF membrane and the chemiluminescent HRP substrate used in Western blotting were obtained from Merck Millipore (Burlington, MA, USA). Flat-bottom microtitration plates were purchased from Corning Inc. (Corning, NY, USA). All other chemicals were of analytical grade and were purchased from Sigma-Aldrich (St. Louis, MO, USA).

Serial dilutions starting at 1:50 of donkey sera were analyzed for Besnoitia antibodies by IFAT using fluorescein (FITC)-conjugate solution [affinity purified goat anti-horse IgG (H + L) (Jackson ImmunoResearch Laboratories, West Grove, USA; diluted 1:50 in PBS)]. The procedure was performed as previously described [10 (link)]. Sera from infected or non-infected donkeys were used as positive and negative controls, respectively, sampled during a besnoitiosis outbreak in north-eastern Pennsylvania [7 (link)].