Epizootic hemorrhagic disease (EHD) of deer is an arthropod-borne Orbivirus that causes infection in wild and domestic ruminants. Infection with EHD virus (EHDV) causes fatal disease, especially in white-tailed deer (Odocoileus virginianus), bighorn sheep (Ovis canadensis), mule deer (Odocoileus hemionus), pronghorn (Antilocapra americana), and possibly black-tailed deer (Odocoileus hemionus columbianus). In domestic ruminants, EHDV may induce clinical and pathologic signs similar to those of bluetongue (BT). However, in North America, natural infections caused by EHDV serotype 1 and serotype 2 have rarely been associated with overt clinical disease in cattle. Laboratory diagnosis is most often based on the detection of anti-EHDV antibodies in serum. Among several serologic assays, agar gel immunodiffusion (AGID) is most widely used but is highly subjective, consumes large quantity of reference reagents, and may be complicated by cross-reactions between EHDV serotypes and other orbiviruses, such as BT viruses (BTVs). As an alternative, enzyme-linked immunosorbent assays (ELISAs), both blocking (bELISA) and competitive (cELISA), have been developed in an attempt to overcome the above problems and to serve as large-scale screening tests with objective interpretation. Recently, a cELISA was developed that utilizes an EHDV group-specific monoclonal antibody (MAb) designated C.31, which is group specific and unreactive with antibodies against BTV. After some modifications, the performance of the this cELISA was compared with that of the standard AGID test for detection of anti-EHDV antibodies using bovine field sera collected in Canada (EHD free) and parts of the USA, where EHD is endemic, and deer sera collected in Georgia (USA). Antigen was prepared from baby hamster kidney cells in serum-free medium and infected with plaque-purified EHDV serotype 1, according to a previously described procedure. Sera from serial or paired blood samples from calves experimentally infected and/or challenged with EHDV serotypes 1 and 2, from a calf experimentally infected with BTV serotype 10, and from several calves exposed to South African BTV serotypes 1-16 and 18-20 and the US BTV serotypes 10, 11, 13, and 17 were used to study seroconversion values. 1-5 To establish negative value, 2 panels totalling 1,194