Bovine herpesvirus 1 (BoHV-1) is an infectious agent of concern in the international export of bovine products; it is endemic in the United States, but it has been eradicated in many countries of the European Union (EU). For export of semen to the EU, accurate assessment of BoHV-1 status of the bull is required and is usually accomplished by measuring the level of antibody to the virus. The gold standard is virus neutralization (VN) using overnight incubation with the virus, a test approved by the World Organization for Animal Health (OIE). Enzyme-linked immunosorbent assay (ELISA) is also approved for international trade. The lone U.S. Department of Agriculture-approved commercial ELISA was compromised with specificity problems, which necessitated the development of a different ELISA. Of 4 monoclonal antibodies evaluated, 1 directed against glycoprotein C of BoHV-1 was found to be the most reliable. One hundred twenty-eight characterized positive samples and 334 negative serum samples were tested. The blocking ELISA showed 97.7% sensitivity and 99.4% specificity as compared with OIE VN. The Wisconsin Veterinary Diagnostic Laboratory ELISA fulfills the OIE requirement for a blocking or competitive ELISA to qualify animals for export to BoHV-1-free countries.