Improving species-specific IDMS: the advantages of triple IDMS

Abstract

Triple isotope dilution mass spectrometry (triple IDMS) has been applied for the first time on protein quantification, especially on transferrin. Transferrin as an acute phase protein is a marker for several inflammation processes in the human body. Therefore, in Germany, the accurate and precise measurement of this important analyte is required. In this work, a new approach to triple IDMS is described and compared to double IDMS. Also, complete uncertainty budgets for both methods were set up to demonstrate the ability of this method to be used as a reference procedure. The relative expanded uncertainty (k=2) for triple IDMS (3.6 %) is smaller than the one for double IDMS (4.0 %). The content of transferrin found in the human serum reference material ERM-DA470k/IFCC ((2.41±0.08) g/kg) with both methods was in good agreement with each other and with the certificate. For triple IDMS ((2.426±0.086) g/kg) and for double IDMS ((2.317±0.092) g/kg), transferrin was determined. Although triple IDMS is a little more time consuming compared to double IDMS, there is the advantage that the isotopic composition of the spike material does not have to be determined. This is very useful especially in case of a marginal isotopic enrichment in the spike or problems with the accurate measurement of the spike isotope ratio.

Measurement Strategies

“…The uncertainty is improved only marginally as compared to double IDMS and is significantly more laborious. 27,28 For more details on the calibration strategies and on the blank calculations the reader is referred to the literature. 15,27,29 Independent of the calibration strategy, a blank of the complete analytical procedure has to be measured and usually the blank can be spiked and treated exactly like a sample.…”

Update on the status of metrology for metalloproteins

“…To reduce the inuence of Fe background and low isotopic enrichment in the spike material, triple isotope dilution mass spectrometry (IDMS) was used for the quantication of total TRF. 19 Besides the determination of total TRF, the quantication of the various sialoforms is important as described above. One method to separate the various sialoforms of TRF in serum is the use of capillary electrophoresis (CE) coupled with UV/Vis or MS detection aer purication of the sample using an immunoaffinity cartridge.…”

Atomic spectrometry updates. Review of advances in elemental speciation

“…Triple isotope dilution mass spectrometry (ID-MS) has been used to quantify transferrin in a human serum CRM, ERM-DA470k/IFCC. 63 The hetero atom used for quantication was Fe, with 57 Fe being the spike isotope and an Fe saturation procedure was used to ensure a uniform Fe/protein stoichiometry. An anion exchange column, Mono-Q, 5/50 GL, with an aqueous based gradient elution was used to separate the transferrin from other matrix components.…”