Suspected bacterial urinary tract infections (UTI) are a common cause of overuse and misuse of antimicrobials. A bedside diagnostic test that could accurately predict urine culture results would prevent antimicrobial overuse, but accurate biomarkers have not yet been identified in veterinary medicine. The objective of this study was to evaluate urine myeloperoxidase (uMPO) as a rapidly available, accurate marker to predict urine culture results. We hypothesized that uMPO would be higher in dogs with a positive urine culture than in dogs with a negative urine culture, and that uMPO could be used to aid in the accurate diagnosis of significant bacteriuria. Urine samples were collected from a veterinary university clinical pathology lab. uMPO concentration was measured using a commercially available canine myeloperoxidase (MPO) enzyme-linked immunosorbent assay (ELISA). Following validation, samples from 98 dogs that had a urinalysis and urine culture performed as part of their diagnostic investigation were included. Forty-seven dogs had a negative urine culture and fifty-one dogs had a positive urine culture. uMPO levels were significantly higher in samples that had a positive culture (median 2.13 ng/ml; IQR 0.98-7.07) versus samples that had a negative culture (median 1.07 ng/ml; IQR 0.52-1.84)(p < 0.005). Based on receiver-operator characteristic, a cutoff of 0.55 ng/ml was chosen to maximize sensitivity and specificity. Using a cutoff of 0.55 ng/ml, uMPO had a sensitivity of 70% and specificity of 69% to determine the presence of a positive culture. However, the degree of overlap between groups may preclude the use of this test as a surrogate for urine culture in a clinical setting.