2.4 Serum bactericidal activity (SBA)
against different strains was measured as follows. The blood samples
collected from clinically healthy horses for routine veterinary control
purposes were used. The samples were collected into a yellow-cap vacuum
tube with clot activator (Elamed, Moscow, Russia). The serum was
separated by centrifugation at 1600 × g for 10 min. The serum was stored
at +4 oC and used for the tests within 24 h. For the
SBA assessment, the wells of 96-well plate containing 175 μl of LB
medium and 25 μl of the serum were inoculated with 5 μl of the
corresponding strain mid-log phase culture (OD600 = 0.6)
and the plates were incubated at 37oC in an automated
plate reader with agitation. The OD600 was recorded
every 30 min. In the control experiment the same volume of physiological
saline replaced the serum. The whole experiment was triplicated.