Absolute id p180 kit

To assess the metabolite profiling, the Absolute ID p180 Kit (Biocrates Life Sciences AG, Innsbruck, Australia) for mass spectrometry was used. The quantification of 95 metabolites was done for 200 participants. Metabolites were composed of 67 Glycerophospholipids (GPs), 12 Acylcarnitines (ACs), 10 Sphingolipids (SGs) and 6 amino acids (AA). For GPs, ACs and SGs x:y notation was used, x denotes the number of carbons in the side chain and y the number of double bonds. The lipids were subdivided into different classes: 28 Phosphatidylcholines diacyl (PCaa); 36 Phosphatidylcholines acyl-alkyl (PCae); 3 LysoPhosphatidylcholines acyl (LysoPC); 4 Hydroxyshingomyelins; and 6 Shingomyelins (SM). GPs are differentiated with respect to the presence of ester (a) and ether (e) bonds in the glycerol moiety. Two letters indicate that the first and second positions of glycerol unit are bound to a fatty acid residue, while a single letter (a or e) indicates a bond with only one fatty acid residue. The concentration of all metabolites was reported in μM. Metabolites for which more than half of the values were below the limit of detection and or with standard out of range were excluded.

Phase extraction of metabolites from the frozen kidney samples of mice was performed as previously described (64 (link)). The AbsoluteID p180 Kit (Biocrates Life Sciences) was used to determine 186 metabolites. The samples were measured on mass spectrometer QTRAP 4500 (Sciex), in combination with a high-performance liquid chromatography (Agilent Technologies). The concentrations of the metabolites were calculated automatically by the MetIDQ software (Biocrates Life Sciences). The levels of reduced glutathione and oxidized glutathione were quantified with a luminescence-based glutathione assay (GSH-Glo; Promega). See Supplemental Table 3 for the summary of all significantly altered metabolites in the kidneys from Stk25^–/– versus WT mice.

As previously described [35 (link)], the Absolute ID p180 Kit (Biocrates Life Sciences AG, Innsbruck, Australia) for mass spectrometry was used for the metabolic profiling measurements for two-hundred participants. Ninety-five metabolites were quantified. They include: 67 Glycerophospholipids (GPs), 12 AC, 10 Sphingolipids (SGs) and 6 AAs. For GPs, ACs and SGs x:y notation was used, x denoting the number of carbons in the side chain and y the number of double bonds. All metabolite concentrations are presented in μM. A metabolite would have been excluded if more than half of the values obtained were below the limit of detection or with standard out of range.