Technical Abstract: Ovine progressive pneumonia virus (OPPV) infects at least one sheep in eighty-one percent of U.S. sheep flocks as measured by serological diagnostic tests and can cause viral-induced mastitis, arthritis, dypsnea, and cachexia. Diagnostic tests that quantify OPP proviral load in peripheral blood leukocytes (PBL) may aid in the identification of infected and super-spreader OPPV sheep and are highly sought. In this study, we developed primers and a Taqman® probe to the highly conserved transmembrane (tm) region of the OPPV envelope gene and established a real-time quantitative (q) PCR assay. This new OPPV qPCR assay was evaluated against a competitive inhibition enzyme-linked immunosorbent assay (cELISA) using 396 PBL samples and sera from Dubois, Idaho sheep ranging in ages 3 to 6 years of the Columbia, Rambouillet and Polypay breeds. The new OPPV qPCR had a positive concordance of 96.2±2.3% and a negative concordance of 97.7±2.5% as compared to the cELISA with a kappa value of 0.93 indicating excellent agreement between the two tests. In addition, the presence of tm was confirmed from the three OPPV qPCR positive and cELISA negative sheep and from fifteen sheep with different OPP proviral loads by cloning and sequencing. These data indicate that the OPPV qPCR may be used as a confirmatory or supplemental diagnostic tool for OPPV infection.