Reproducibility of Targeted Lipidome Analyses (Lipidyzer) in Plasma and Erythrocytes over a 6-Week Period

Abstract

It is essential to measure lipid biomarkers with a high reproducibility to prevent biased results. We compared the lipid composition and inter-day reproducibility of lipid measurements in plasma and erythrocytes. Samples from 42 individuals (77% women, mean age 65 years, mean body mass index (BMI) 27 kg/m2), obtained non-fasted at baseline and after 6 weeks, were used for quantification of up to 1000 lipid species across 13 lipid classes with the Lipidyzer platform. Intraclass correlation coefficients (ICCs) were calculated to investigate the variability of lipid concentrations between timepoints. The ICC distribution of lipids in plasma and erythrocytes were compared using Wilcoxon tests. After data processing, the analyses included 630 lipids in plasma and 286 in erythrocytes. From these, 230 lipids overlapped between sample types. In plasma, 78% of lipid measurements were reproduced well to excellently, compared to 37% in erythrocytes. The ICC score distribution in plasma (median ICC 0.69) was significantly better than in erythrocytes (median ICC 0.51) (p-value < 0.001). At the class level, reproducibility in plasma was superior for triacylglycerols and cholesteryl esters while ceramides, diacylglycerols, (lyso)phosphatidylethanolamines, and sphingomyelins showed better reproducibility in erythrocytes. Although in plasma overall reproducibility was superior, differences at individual and class levels may favor the use of erythrocytes.

Cross-Laboratory Standardization of Preclinical Lipidomics Using Differential Mobility Spectrometry and Multiple Reaction Monitoring

“…Considering these circumstances, three sites robustly measured more than 500 lipid species in the cohort plasma samples. A recent study on reproducibility of targeted lipidome analyses in plasma and erythrocytes using the Lipidyzer platform also reported that 491 lipid measurements in plasma were reproduced well over a 6 week period . This promising number of robustly measured lipids puts the platform in a strong position to be used in important clinical research and biomarker discovery studies.…”

“…A recent study on reproducibility of targeted lipidome analyses in plasma and erythrocytes using the Lipidyzer platform also reported that 491 lipid measurements in plasma were reproduced well over a 6 week period. 32 This promising number of robustly measured lipids puts the platform in a strong position to be used in important clinical research and biomarker discovery studies. At present, it is still a challenging task to reproduce clinical metabolomics findings between laboratories, with the major impediment being the limited methodological and practical agreement among laboratories.…”

Sphingolipidome Quantification by Liquid Chromatography- High Resolution Mass Spectrometry: Whole Blood vs. Plasma

“…Recent results suggested that sphingolipids overload in RBCs could occur during erythropoiesis [41]. Also as recently reported that differences at individual levels in plasma may favor the use of erythrocytes [42] our study investigated the best protocol to do that by using the minimum volume of whole blood and the relevant internal standards for normalization. We will move next to a case-control study of a particular disease to show the applicability of this new approach.…”

Analytical Considerations for Reducing the Matrix Effect for the Sphingolipidome Quantification in Whole Blood

“…The LOQ for all the sphingolipids investigated here allows the use of only 10-μl WB. A recent study looking at the basal lipids level in healthy volunteers over a 6 weeks period in plasma and erythrocytes reported that reproducibility in ceramides, diacylglycerols and sphingomyelins was better in erythrocytes [ 39 ]. To reduce any artifacts that could results from erythrocytes isolation, our study investigated the best protocol to use WB, by using the minimum volume necessary for accurate quantification and the relevant ISs for normalization.…”