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von Toerne, C. ; Kahle-Stephan, M. ; Schäfer, A. ; Ispiryan, R. ; Blindert, M. ; Hrabě de Angelis, M. ; Neschen, S. ; Ueffing, M. ; Hauck, S.M.

Apoe, Mbl2 and Psp plasma protein levels correlate with diabetic phenotype in NZO mice - an optimized rapid workflow for SRM-based quantification.

J. Proteome Res. 12, 1331-1343 (2013)

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Open Access Green as soon as Postprint is submitted to ZB.

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Male New Zealand Obese (NZO) mice progress through pathophysiological stages similar to humans developing obesity-associated type 2 diabetes (T2D). The current challenge is to establish quantitative proteomics from small plasma sample amounts. We established an analytical workflow that facilitates a reproducible depletion of high-abundance proteins, has high throughput applicability, and allows absolute quantification of proteins from mouse plasma samples by LC-SRM-MS. The ProteoMiner equalizing technology was adjusted to the small sample amount and reproducibility of the identifications was monitored by spike proteins. Based on the label-free relative quantification of proteins in depleted plasma of a test set of NZO mice, assays for potential candidates were designed for the setup of a targeted selected reaction monitoring (SRM) approach and absolute quantification. We could demonstrate that apolipoprotein E (Apoe), mannose-binding lectin 2 (Mbl2) and parotid secretory protein (Psp) are present at significantly different quantities in depleted plasma of diabetic NZO mice compared to non-diabetic controls using AQUA peptides. Quantification was validated for Mbl2 using the ELISA technology on non-depleted plasma. We conclude that the depletion technique is applicable to restricted sample amounts and suitable for the identification of T2D signatures in plasma.

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