Interleukin-10 (IL-10) interferes with monocyte and macrophage activation of Th1 helper lymphocyte production of nitric oxide and synthesis of various inflammatory mediators. An IL-10 homologue (vIL-10) is produced by several herpes viruses and is hypothesized to help the virus down regulate, and thus evade, host immune responses. The gammaherpes rhadinovirus Ovine herpesvirus-2 (OvHV-2) encodes an IL-10 like molecule highly homologous to mammalian IL-10. This virus causes sheep-associated malignant catarrhal fever (MCF), the most common form of MCF in the United States. MCF is a lymphoproliferative and inflammatory syndrome that has delayed clinical presentation hypothesized to be influenced by the production of vIL-10. For this study, the gene encoding vIL-10, Ov2.5 ORF, was amplified by PCR, cloned, sequenced, and a predicted 30 amino acid segment from the amino terminus synthesized. This synthetic peptide was used to develop a novel direct enzyme-linked immunosorbant assay (ELISA) to detect isotype-specific antibodies to OvHV-2 vIL-10. Work to date indicates that lambs do not have detectable levels of maternally derived antibody to vIL-10 during the first few weeks of age. Ewes, which are refractory to clinical infection, generally have high levels of antibody to vIL10. A weak correlation exists between the vIL-10 ELISA and a commercially available competitive-inhibition ELISA (cELISA). We believe the vIL-10 ELISA will refine the ability to identify ruminants exposed to sheepassociated MCF, provide an important tool for determining the role of vIL10 in disease pathogenesis, and may contribute toward the development of a new vaccine strategy for the control of malignant catarrhal fever.