Objectives The aim of this study was to determine the accuracy of a commercial luteinizing hormone (LH) test as an aid in distinguishing between sexually intact and ovariectomized or castrated domestic cats. Methods Convenience serum samples collected from sexually intact female and male cats (n = 67) undergoing elective sterilization surgery and archived sera from ovariectomized and castrated cats (n = 54) were tested for LH using a commercial diagnostic assay. Test results were compared with the known reproductive status of the cats. Additionally, sera from sexually intact (n = 54) and ovariectomized (n = 94) queens were collected at specific times of the year to evaluate possible seasonal effects on test results. Results Overall test sensitivity was 89.3% (95% confidence interval [CI] 82.3-94.2%), specificity was 92.6% (95% CI 87.1-96.2%) and accuracy was 91.1%. Analysis of results of female cats (n = 216) - sexually intact (n = 87) and ovariectomized (n = 129) - yielded a test sensitivity of 90.8% (95% CI 82.7-96.0%), a specificity of 92.3% (95% CI 86.2-96.2%) and accuracy of 91.7%. Analysis of the results of male cats (n = 53) - sexually intact (n = 19) and neutered (n = 34) - yielded test a sensitivity of 85.3% (95% CI 68.9-95.1%), a specificity of 94.7% (95% CI 74.0-99.9%) and accuracy of 88.7%. The sera of 10 intact queens unexpectedly yielded positive LH results; two of these cats were in estrus, based on visual inspection at the time of ovariohysterectomy. Test accuracy was 94.6% for those 148 samples collected at specific times of the year, with two samples each over three, 3 month periods yielding false-positive results. Conclusions and relevance The commercial point-of-care LH test is a useful adjunct to historical and physical examination findings for determination of reproductive status in domestic cats. Repeat testing 24 h later should be considered for those female cats with signs of estrus and initial positive test results.