The study aimed to evaluate the usability of sterile bag collection (SBC) urinalysis and urine culture for diagnosing urinary tract infections (UTI). Urine culture is key for diagnosing UTI, and transurethral catheterization (TUC) or suprapubic aspiration is recommended for non-toilet-trained children. Although urine testing using SBC is non-invasive and easy, UTI can be diagnosed only if other criteria including clinical symptoms and positive urinalysis results are met. This study included 228 infants who were hospitalized for unexplained fever from October 2015 to June 2016. TUC culture, SBC urinalysis, and urine culture were performed for all patients. UTI was diagnosed when the TUC culture results met the criterion of ≥104 colony-forming units (CFU)/mL. When UTI diagnosis was made based on SBC urine colony counts ≥105 CFU/mL, the false-positive and false-negative rates were 6.3% and 70.0%, respectively. When the criterion was set as ≥104 CFU/mL, they were 23.7% and 30.0%, respectively. When both the criteria of ≥105 CFU/mL and positive urinalysis results were met, the false-positive rate was 2.4%, and the false-negative rate was 80%. Our results suggest that diagnosing UTI using SBC urinalysis and urine culture is not useful in infants with unexplained fever.