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Description
Exposure of blood plasma/serum (P/S) to thawed conditions, greater than -30°C, can produce biomolecular changes that misleadingly impact measurements of clinical markers within archived samples. Reported here is a low sample-volume, dilute-and-shoot, intact protein mass spectrometric assay of albumin proteoforms called “ΔS-Cys-Albumin” that quantifies cumulative exposure of archived P/S samples to thawed conditions. The assay uses the fact that S-cysteinylation (oxidation) of albumin in P/S increases to a maximum value when exposed to temperatures greater than -30°C. The multi-reaction rate law that governs this albumin S-cysteinylation formation in P/S was determined and was shown to predict the rate of formation of S-cysteinylated albumin in P/S samples—a step that enables back-calculation of the time at which unknown P/S specimens have been exposed to room temperature. To emphasize the capability of this assay, a blind challenge demonstrated the ability of ΔS-Cys-Albumin to detect exposure of individual and grouped P/S samples to unfavorable storage conditions. The assay was also capable of detecting an anomaly in a case study of nominally pristine serum samples collected under NIH-sponsorship, demonstrating that empirical evidence is required to guarantee accurate knowledge of archived P/S biospecimen storage history.
The ex vivo glycation of human serum albumin was also investigated showing that P/S samples stored above their freezing point leads to significant increases in glycated albumin. These increases were found to occur within hours at room temperature, and within days at -20 °C. These increases continued over a period of 1-2 weeks at room temperature and over 200 days at -20 °C, ultimately resulting in a doubling of glycated albumin in both healthy and diabetic patients. It was also shown that samples stored at lower surface area-to-volume ratios or incubated under a nitrogen atmosphere experienced less rapid glucose adduction of albumin—suggesting a role for oxidative glycation in the ex vivo glycation of albumin.
The ex vivo glycation of human serum albumin was also investigated showing that P/S samples stored above their freezing point leads to significant increases in glycated albumin. These increases were found to occur within hours at room temperature, and within days at -20 °C. These increases continued over a period of 1-2 weeks at room temperature and over 200 days at -20 °C, ultimately resulting in a doubling of glycated albumin in both healthy and diabetic patients. It was also shown that samples stored at lower surface area-to-volume ratios or incubated under a nitrogen atmosphere experienced less rapid glucose adduction of albumin—suggesting a role for oxidative glycation in the ex vivo glycation of albumin.
ContributorsJeffs, Joshua W (Author) / Borges, Chad R (Thesis advisor) / Van Horn, Wade (Committee member) / Williams, Peter (Committee member) / Arizona State University (Publisher)
Created2018