Effects of sample handling and storage on quantitative lipid analysis in human serum

Angela M. Zivkovic, Michelle M. Wiest, Uyen Thao Nguyen, Ryan Davis, Steven M. Watkins, J. Bruce German

Research output: Contribution to journalArticle

84 Scopus citations

Abstract

There is sparse information about specific storage and handling protocols that minimize analytical error and variability in samples evaluated by targeted metabolomics. Variance components that affect quantitative lipid analysis in a set of human serum samples were determined. The effects of freeze-thaw, extraction state, storage temperature, and freeze-thaw prior to density-based lipoprotein fractionation were quantified. The quantification of high abundance metabolites, representing the biologically relevant lipid species in humans, was highly repeatable (with coefficients of variation as low as 0.01 and 0.02) and largely unaffected by 1-3 freeze-thaw cycles (with 0-8% of metabolites affected in each lipid class). Extraction state had effects on total lipid class amounts, including decreased diacylglycerol and increased phosphatidylethanolamine in thawed compared with frozen samples. The effects of storage temperature over 1 week were minimal, with 0-4% of metabolites affected by storage at 4°C, -20°C, or -80°C in most lipid classes, and 19% of metabolites in diacylglycerol affected by storage at -20°C. Freezing prior to lipoprotein fractionation by density ultracentrifugation decreased HDL free cholesterol by 37% and VLDL free fatty acid by 36%, and increased LDL cholesterol ester by 35% compared with fresh samples. These findings suggest that density-based fractionation should preferably be undertaken in fresh serum samples because up to 37% variability in HDL and LDL cholesterol could result from a single freeze-thaw cycle. Conversely, quantitative lipid analysis within unfractionated serum is minimally affected even with repeated freeze-thaw cycles.

Original languageEnglish (US)
Pages (from-to)507-516
Number of pages10
JournalMetabolomics
Volume5
Issue number4
DOIs
StatePublished - Dec 2009

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Keywords

  • Effects of sample handling on analytical accuracy
  • Freeze-thaw
  • Lipoprotein separation
  • Metabolomics
  • Targeted metabolomic lipid analysis

ASJC Scopus subject areas

  • Biochemistry
  • Clinical Biochemistry
  • Endocrinology, Diabetes and Metabolism

Cite this

Zivkovic, A. M., Wiest, M. M., Nguyen, U. T., Davis, R., Watkins, S. M., & German, J. B. (2009). Effects of sample handling and storage on quantitative lipid analysis in human serum. Metabolomics, 5(4), 507-516. https://doi.org/10.1007/s11306-009-0174-2