BACKGROUND: Formal validation of methods for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. A protocol for processing of a biospecimen (urine) was validated for fitness-for-purpose in terms of key downstream endpoints. METHODS: Urine processing was optimized for centrifugation conditions on the basis of microparticle counts at room temperature (RT) and at 4°C. The optimal protocol was validated for performance (microparticle counts), and for reproducibility and robustness for centrifugation temperature (4°C vs. RT) and brake speed (soft, medium, hard). Acceptance criteria were based on microparticle counts, cystatin C and creatinine concentrations, and the metabolomic profile. RESULTS: The optimal protocol was a 20-min, 12,000 g centrifugation at 4°C, and was validated for urine collection in terms of microparticle counts. All reproducibility acceptance criteria were met. The protocol was robust for centrifugation at 4°C versus RT for all parameters. The protocol was considered robust overall in terms of brake speeds, although a hard brake gave significantly fewer microparticles than a soft brake. CONCLUSIONS: We validated a urine processing method suitable for downstream proteomic and metabolomic applications. Temperature and brake speed can influence analytic results, with 4°C and high brake speed considered optimal. Laboratories and biobanks should ensure these conditions are systematically recorded in the scope of accreditation.
BACKGROUND: Formal validation of methods for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. A protocol for processing of a biospecimen (urine) was validated for fitness-for-purpose in terms of key downstream endpoints. METHODS: Urine processing was optimized for centrifugation conditions on the basis of microparticle counts at room temperature (RT) and at 4°C. The optimal protocol was validated for performance (microparticle counts), and for reproducibility and robustness for centrifugation temperature (4°C vs. RT) and brake speed (soft, medium, hard). Acceptance criteria were based on microparticle counts, cystatin C and creatinine concentrations, and the metabolomic profile. RESULTS: The optimal protocol was a 20-min, 12,000 g centrifugation at 4°C, and was validated for urine collection in terms of microparticle counts. All reproducibility acceptance criteria were met. The protocol was robust for centrifugation at 4°C versus RT for all parameters. The protocol was considered robust overall in terms of brake speeds, although a hard brake gave significantly fewer microparticles than a soft brake. CONCLUSIONS: We validated a urine processing method suitable for downstream proteomic and metabolomic applications. Temperature and brake speed can influence analytic results, with 4°C and high brake speed considered optimal. Laboratories and biobanks should ensure these conditions are systematically recorded in the scope of accreditation.
Authors: Uta Erdbrügger; Charles J Blijdorp; Irene V Bijnsdorp; Francesc E Borràs; Dylan Burger; Benedetta Bussolati; James Brian Byrd; Aled Clayton; James W Dear; Juan M Falcón-Pérez; Cristina Grange; Andrew F Hill; Harry Holthöfer; Ewout J Hoorn; Guido Jenster; Connie R Jimenez; Kerstin Junker; John Klein; Mark A Knepper; Erik H Koritzinsky; James M Luther; Metka Lenassi; Janne Leivo; Inge Mertens; Luca Musante; Eline Oeyen; Maija Puhka; Martin E van Royen; Catherine Sánchez; Carolina Soekmadji; Visith Thongboonkerd; Volkert van Steijn; Gerald Verhaegh; Jason P Webber; Kenneth Witwer; Peter S T Yuen; Lei Zheng; Alicia Llorente; Elena S Martens-Uzunova Journal: J Extracell Vesicles Date: 2021-05-21
Authors: Pauline Erpicum; Oriane Hanssen; Laurent Weekers; Pierre Lovinfosse; Paul Meunier; Luaba Tshibanda; Jean-Marie Krzesinski; Roland Hustinx; François Jouret Journal: Clin Kidney J Date: 2016-09-06