Chirag R Parikh1, Isabel Butrymowicz2, Angela Yu2, Vernon M Chinchilli3, Meyeon Park4, Chi-Yuan Hsu5, W Brian Reeves6, Prasad Devarajan7, Paul L Kimmel8, Edward D Siew9, Kathleen D Liu4. 1. Section of Nephrology, Department of Medicine, Yale University School of Medicine, New Haven, CT; Program of Applied Translational Research, Yale University School of Medicine, New Haven, CT. Electronic address: chirag.parikh@yale.edu. 2. Section of Nephrology, Department of Medicine, Yale University School of Medicine, New Haven, CT; Program of Applied Translational Research, Yale University School of Medicine, New Haven, CT. 3. Department of Public Health Sciences, Penn State College of Medicine, Hershey, PA. 4. Division of Nephrology, Department of Medicine, University of California, San Francisco, CA. 5. Division of Nephrology, Department of Medicine, University of California, San Francisco, CA; Division of Research, Kaiser Permanente of Northern California, Oakland, CA. 6. Department of Medicine, Division of Nephrology, Penn State College of Medicine, Hershey, PA. 7. Nephrology and Hypertension, Cincinnati Children's Hospital Medical Center, Cincinnati, OH. 8. Division of Kidney, Urologic, and Hematologic Diseases, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD. 9. Department of Medicine, Division of Nephrology and Hypertension, Vanderbilt University Medical Center, Nashville, TN.
Abstract
BACKGROUND: The study of novel urinary biomarkers of acute kidney injury has expanded exponentially. Effective interpretation of data and meaningful comparisons between studies require awareness of factors that can adversely affect measurement. We examined how variations in short-term storage and processing might affect the measurement of urine biomarkers. STUDY DESIGN: Cross-sectional prospective. SETTING & PARTICIPANTS: Hospitalized patients from 2 sites: Yale New Haven Hospital (n=50) and University of California, San Francisco Medical Center (n=36). PREDICTORS: We tested the impact of 3 urine processing conditions on these biomarkers: (1) centrifugation and storage at 4°C for 48 hours before freezing at -80°C, (2) centrifugation and storage at 25°C for 48 hours before freezing at -80°C, and (3) uncentrifuged samples immediately frozen at -80°C. OUTCOMES: Urine concentrations of 5 biomarkers: neutrophil gelatinase-associated lipocalin (NGAL), interleukin 18 (IL-18), kidney injury molecule 1 (KIM-1), liver-type fatty acid-binding protein (L-FABP), and cystatin C. MEASUREMENTS: We measured urine biomarkers by established enzyme-linked immunosorbent assay methods. Biomarker values were log-transformed, and agreement with a reference standard of immediate centrifugation and storage at -80°C was compared using concordance correlation coefficients (CCCs). RESULTS: Neither storing samples at 4°C for 48 hours nor centrifugation had a significant effect on measured levels, with CCCs higher than 0.9 for all biomarkers tested. For samples stored at 25°C for 48 hours, excellent CCC values (>0.9) also were noted between the test sample and the reference standard for NGAL, cystatin C, L-FABP and KIM-1. However, the CCC for IL-18 between samples stored at 25°C for 48 hours and the reference standard was 0.81 (95% CI, 0.66-0.96). LIMITATIONS: No comparisons to fresh, unfrozen samples; no evaluation of the effect of protease inhibitors. CONCLUSIONS: All candidate markers tested using the specified assays showed high stability with both short-term storage at 4°C and without centrifugation prior to freezing. For optimal fidelity, urine for IL-18 measurement should not be stored at 25°C before long-term storage or analysis.
BACKGROUND: The study of novel urinary biomarkers of acute kidney injury has expanded exponentially. Effective interpretation of data and meaningful comparisons between studies require awareness of factors that can adversely affect measurement. We examined how variations in short-term storage and processing might affect the measurement of urine biomarkers. STUDY DESIGN: Cross-sectional prospective. SETTING & PARTICIPANTS: Hospitalized patients from 2 sites: Yale New Haven Hospital (n=50) and University of California, San Francisco Medical Center (n=36). PREDICTORS: We tested the impact of 3 urine processing conditions on these biomarkers: (1) centrifugation and storage at 4°C for 48 hours before freezing at -80°C, (2) centrifugation and storage at 25°C for 48 hours before freezing at -80°C, and (3) uncentrifuged samples immediately frozen at -80°C. OUTCOMES: Urine concentrations of 5 biomarkers: neutrophil gelatinase-associated lipocalin (NGAL), interleukin 18 (IL-18), kidney injury molecule 1 (KIM-1), liver-type fatty acid-binding protein (L-FABP), and cystatin C. MEASUREMENTS: We measured urine biomarkers by established enzyme-linked immunosorbent assay methods. Biomarker values were log-transformed, and agreement with a reference standard of immediate centrifugation and storage at -80°C was compared using concordance correlation coefficients (CCCs). RESULTS: Neither storing samples at 4°C for 48 hours nor centrifugation had a significant effect on measured levels, with CCCs higher than 0.9 for all biomarkers tested. For samples stored at 25°C for 48 hours, excellent CCC values (>0.9) also were noted between the test sample and the reference standard for NGAL, cystatin C, L-FABP and KIM-1. However, the CCC for IL-18 between samples stored at 25°C for 48 hours and the reference standard was 0.81 (95% CI, 0.66-0.96). LIMITATIONS: No comparisons to fresh, unfrozen samples; no evaluation of the effect of protease inhibitors. CONCLUSIONS: All candidate markers tested using the specified assays showed high stability with both short-term storage at 4°C and without centrifugation prior to freezing. For optimal fidelity, urine for IL-18 measurement should not be stored at 25°C before long-term storage or analysis.
Authors: Ashish Agrawal; Michael A Matthay; Kirsten N Kangelaris; John Stein; Jeffrey C Chu; Brandon M Imp; Alfredo Cortez; Jason Abbott; Kathleen D Liu; Carolyn S Calfee Journal: Am J Respir Crit Care Med Date: 2013-04-01 Impact factor: 21.405
Authors: Michael Bennett; Catherine L Dent; Qing Ma; Sudha Dastrala; Frank Grenier; Ryan Workman; Hina Syed; Salman Ali; Jonathan Barasch; Prasad Devarajan Journal: Clin J Am Soc Nephrol Date: 2008-03-12 Impact factor: 8.237
Authors: Chirag R Parikh; Alkesh Jani; Vyacheslav Y Melnikov; Sarah Faubel; Charles L Edelstein Journal: Am J Kidney Dis Date: 2004-03 Impact factor: 8.860
Authors: Kathleen D Liu; Wei Yang; Amanda H Anderson; Harold I Feldman; Sevag Demirjian; Takayuki Hamano; Jiang He; James Lash; Eva Lustigova; Sylvia E Rosas; Michael S Simonson; Kaixiang Tao; Chi-yuan Hsu Journal: Kidney Int Date: 2013-01-23 Impact factor: 10.612
Authors: Krista M Gibbs; Jenelle M Izer; W Brian Reeves; Ronald P Wilson; Timothy K Cooper Journal: J Am Assoc Lab Anim Sci Date: 2018-12-11 Impact factor: 1.232
Authors: Sushrut S Waikar; Venkata Sabbisetti; Johan Ärnlöv; Axel C Carlsson; Josef Coresh; Harold I Feldman; Meredith C Foster; Gudeta D Fufaa; Johanna Helmersson-Karlqvist; Chi-Yuan Hsu; Paul L Kimmel; Anders Larsson; Yumin Liu; Lars Lind; Kathleen D Liu; Theodore E Mifflin; Robert G Nelson; Ulf Risérus; Ramachandran S Vasan; Dawei Xie; Xiaoming Zhang; Joseph V Bonventre Journal: Nephrol Dial Transplant Date: 2016-06-07 Impact factor: 5.992
Authors: Meredith P Schuh; Edward Nehus; Qing Ma; Christopher Haffner; Michael Bennett; Catherine D Krawczeski; Prasad Devarajan Journal: Am J Kidney Dis Date: 2015-05-29 Impact factor: 8.860
Authors: Annegret Ulke-Lemée; Arthur Lau; Michelle C Nelson; Matthew T James; Daniel A Muruve; Justin A MacDonald Journal: Inflammation Date: 2018-08 Impact factor: 4.092
Authors: Michelle C Starr; David J Askenazi; Stuart L Goldstein; James W MacDonald; Theo K Bammler; Zahra Afsharinejad; Patrick D Brophy; Sandra E Juul; Dennis E Mayock; Sangeeta R Hingorani Journal: Pediatr Nephrol Date: 2017-08-19 Impact factor: 3.714
Authors: Peter P Reese; Isaac E Hall; Francis L Weng; Bernd Schröppel; Mona D Doshi; Rick D Hasz; Heather Thiessen-Philbrook; Joseph Ficek; Veena Rao; Patrick Murray; Haiqun Lin; Chirag R Parikh Journal: J Am Soc Nephrol Date: 2015-09-15 Impact factor: 10.121
Authors: Jennifer Palermo; Allison B Dart; Alanna De Mello; Prasad Devarajan; Ronald Gottesman; Gonzalo Garcia Guerra; Greg Hansen; Ari R Joffe; Cherry Mammen; Nick Majesic; Catherine Morgan; Peter Skippen; Michael Pizzi; Ana Palijan; Michael Zappitelli Journal: Pediatr Crit Care Med Date: 2017-06 Impact factor: 3.624