BACKGROUND: Hematuria is the most frequent presenting symptom in the vast majority of bladder cancer (BC) patients. The current recommended evaluation of hematuria includes cross sectional imaging and cystoscopy with possible high negative results, expensive costs and substantial patient burden. OBJECTIVES: To validate novel urine-based mRNA-dependant tests; Xpert test and urinary metabolomics assay (CRAT and SLC 25A20genes expression) for BC detection in patients with hematuria. METHODS: Patients presented with hematuria to our tertiary care hospital were evaluated by CT urogram and office white light cystoscopy with subsequent inpatient biopsy for positive findings. Voided precystoscopy urine samples were prospectively collected. Xpert test, assay of targeted urinary metabolomics and cytology, were performed. The tests characteristics presumably were calculated based on the ability to identify BC noninvasively. RESULTS: Between March 2018 and June 2019, 181 patients were included in the final analysis with mean (±SD) age 62 (±10) years with 168 (92.8%) males. Macroscopic hematuria was encountered in 153 (84.5%) patients with irritative bladder symptoms in 48 (26.5%) patients. BC was confirmed by cystoscopy/biopsy in 36 (19.9%) patients. The performance characteristics of Xpert alone (SN: 73%, SP: 83%, NPV: 92%, PPV: 52%) (AUC 0.84, 95% CI 0.75–0.93, p = 0.001), metabolomics assay alone (SN: 89%, SP: 93%, NPV: 97%, PPV: 78%) (AUC 0.91, 95% CI 0.85–0.98, p < 0.001) and combination of both test results (SN: 66%, SP: 98%, NPV: 92%, PPV: 97%) (AUC 0.83, 95% CI 0.74–0.93, p = 0.001) were notably superior to urine cytology (SN: 30%, SP: 84%, NPV: 83%, PPV: 33%) (AUC 0.58, 95% CI 0.47–0.69, p = 0.154) for BC prediction. Cystoscopy-negative patients (CNP) were followed-up for a median (range) 12 (2–19) months. Re-cystoscopy was done for 35 patients with persistent symptoms. BC was diagnosed in 6 patients. Xpert and urinary metabolomics results were observably positive in those 6 patients. CONCLUSION: Xpert test and assay of urinary metabolomics (CRAT and SLC 25A20 genes expression) have the potential for BC detection in hematuria patients. These non invasive urine based tests can help prioritization of the use of invasive diagnostic tests in systems with long waiting times.