title: Highly sensitive reversed-phase high-performance liquid chromatography assay for the detection of Tamm-Horsfall protein in human urine.
abstract: BACKGROUND
  Tamm-Horsfall protein (also known as uromodulin) is the most abundant urinary protein in healthy individuals. Since initially characterized by Tamm and Horsfall, the amount of urinary excretion and structural mutations of Tamm-Horsfall protein is associated with kidney diseases. However, currently available assays for Tamm-Horsfall protein, which are mainly enzyme-linked immunosorbent assay-based, suffer from poor reproducibility and might give false negative results.
  
  
  METHODS
  We developed a novel, quantitative assay for Tamm-Horsfall protein using reversed-phase high-performance liquid chromatography. A precipitation pretreatment avoided urine matrix interference and excessive sample dilution. High-performance liquid chromatography optimization based on polarity allowed excellent separation of Tamm-Horsfall protein from other major urine components.
  
  
  RESULTS
  Our method exhibited high precision (based on the relative standard deviations of intraday [≤2.77%] and interday [≤5.35%] repetitions). The Tamm-Horsfall protein recovery rate was 100.0-104.2%. The mean Tamm-Horsfall protein concentration in 25 healthy individuals was 31.6 ± 18.8 mg/g creatinine. There was a strong correlation between data obtained by high-performance liquid chromatography and enzyme-linked immunosorbent assay (r = 0.906), but enzyme-linked immunosorbent assay values tended to be lower than high-performance liquid chromatography values at low Tamm-Horsfall protein concentrations.
  
  
  CONCLUSIONS
  The high sensitivity and reproducibility of our Tamm-Horsfall protein assay will reduce the number of false negative results of the sample compared with enzyme-linked immunosorbent assay. Moreover, our method is superior to other high-performance liquid chromatography methods, and a simple protocol will facilitate further research on the physiological role of Tamm-Horsfall protein.
authors.affiliation: Clinical Chemistry, Division of Biochemical Science and Technology, Department of Health Sciences, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Masaru
authors.initials: M
authors.lastname: Akimoto
authors.affiliation: Clinical Chemistry, Division of Biochemical Science and Technology, Department of Health Sciences, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Eisaku
authors.initials: E
authors.lastname: Hokazono
authors.affiliation: Clinical Chemistry, Division of Biochemical Science and Technology, Department of Health Sciences, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan.
authors.forename: Eri
authors.initials: E
authors.lastname: Ota
authors.affiliation: Clinical Chemistry, Division of Biochemical Science and Technology, Department of Health Sciences, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan Department of Medical Laboratory Science, Faculty of Health Sciences, Junshin Gakuen University, Fukuoka, Japan.
authors.forename: Takiko
authors.initials: T
authors.lastname: Tateishi
authors.affiliation: Clinical Chemistry, Division of Biochemical Science and Technology, Department of Health Sciences, Faculty of Medical Sciences, Kyushu University, Fukuoka, Japan kayamori@hs.med.kyushu-u.ac.jp.
authors.forename: Yuzo
authors.initials: Y
authors.lastname: Kayamori
mesh_terms: D053000:Analytic Sample Preparation Methods; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D056148:Chromatography, Reverse-Phase / Q000379:methods*; D006801:Humans; D057230:Limit of Detection; D015203:Reproducibility of Results; D012492:Salts / Q000737:chemistry; D016482:Urinalysis / Q000379:methods*; D058949:Uromodulin / Q000302:isolation & purification / Q000652:urine*
chemical_list: D012492:Salts; D058949:Uromodulin
keywords: Tamm–Horsfall protein; enzyme-linked immunosorbent assay; reversed-phase high-performance liquid chromatography; uromodulin