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Title

Method Validation for Preparing Serum and Plasma Samples from Human Blood for Downstream Proteomic, Metabolomic, and Circulating Nucleic Acid-Based Applications
Authors
Ammerlaan, Wim
Trezzi, Jean-Pierre
Lescuyer, Pierre
Mathay, Conny
Hiller, Karsten
Betsou, Fay
Published in Biopreservation and Biobanking. 2014
Abstract Background: Formal method validation for biospecimen processing in the context of accreditation in laboratories and biobanks is lacking. Serum and plasma processing protocols were validated for fitness-for-purpose in terms of key downstream endpoints, and this article demonstrates methodology for biospecimen processing method validation. Methods: Serum and plasma preparation from human blood was optimized for centrifugation conditions with respect to microparticle counts. Optimal protocols were validated for methodology and reproducibility in terms of acceptance criteria based on microparticle counts, DNA and hemoglobin concentration, and metabolomic and proteomic profiles. These parameters were also used to evaluate robustness for centrifugation temperature (4°C versus room temperature [RT]), deceleration (low, medium, high) and blood stability (after a 2-hour delay). Results: Optimal protocols were 10-min centrifugation for serum and 20-min for plasma at 2000 g, medium brake, RT. Methodology and reproducibility acceptance criteria were met for both protocols except for reproducibility of plasma metabolomics. Overall, neither protocol was robust for centrifugation at 4°C versus RT. RT gave higher microparticles and free DNA yields in serum, and fewer microparticles with less hemolysis in plasma. Overall, both protocols were robust for fast, medium, and low deceleration, with a medium brake considered optimal. Pre-centrifugation stability after a 2-hour delay was seen at both temperatures for hemoglobin concentration and proteomics, but not for microparticle counts. Conclusions: We validated serum and plasma collection methods suitable for downstream protein, metabolite, or free nucleic acid-based applications. Temperature and pre-centrifugation delay can influence analytic results, and laboratories and biobanks should systematically record these conditions in the scope of accreditation.
Identifiers
PMID: 25075813
Full text
Article (Author postprint) (296 Kb) - document accessible for UNIGE members only Limited access to UNIGE
Structures
Faculté de médecine / Section de médecine fondamentale / Département de biologie structurale et bioinformatique
Research group Chimie et protéomique clinique (270)
Citation
(ISO format) 		AMMERLAAN, Wim et al. Method Validation for Preparing Serum and Plasma Samples from Human Blood for Downstream Proteomic, Metabolomic, and Circulating Nucleic Acid-Based Applications. In: Biopreservation and Biobanking, 2014. https://archive-ouverte.unige.ch/unige:40614

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Deposited on : 2014-09-30

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