Tamm-Horsfall protein (THP) is the most abundant protein in the urine. THP is expressed in the renal tubule as a precursor protein having 640 amino acid residues and anchored by GPI anchor in the cell membrane. Thereafter, THP is secreted as a glycoprotein is a molecular weight of about 95-100 kD. Despite several investigations from the perspective of renal failure, the physiological role of this protein is not yet clear. It has been reported that THP is also related to certain conditions, such as kidney stone formation and urinary tract infection. To examine the excretion of THP into urine, we constructed an enzyme linked immunosorbent assay(ELISA) for the measurement of THP in the urine. THP was purified from healthy human urine using Diatomaceous Earth. Then we obtained an antibody to purified THP and constructed a sandwich ELISA assay system to test urine samples. The sensitivity of measurement was 0.78 ng/ml. In this assay, the concentration of THP in spot urine can be linearly measured from 0.78 ng/ml to 50 ng/ml. The CV of assay was 2.4 to 4.1%. The measurement was not disrupted by urinary albumin (approximately 15 mg/ml), hemoglobin (approximately 15 microg/ml), glucose (approximately 30 mg/ml) and ascorbic acid (approximately 10 mg/ml). Pretreatment by centrifugation or filtration of urine affected the concentration of THP because of the agglutinated form of THP. We showed that the urinary excretion rate remained almost constant in our test population, 1.00-1.65 mg/hr (average +/- 1SD 1.30 +/- 0.25) for healthy men and 0.61-1.51 mg/hr (0.90 +/- 0.30) for healthy women and 1.10 +/- 0.34 mg/hr overall.