We have investigated species-specific isotope dilution mass spectrometry (IDMS) for the quantification of ferritin-bound iron in murine serum and brain. Therefore, fresh samples were analyzed using size exclusion chromatography inductively coupled plasma mass spectrometry (SEC-ICP-MS). Isotopically labeled (57Fe)ferritin was used as calibrant for the quantification of ferritin-bound iron in murine samples. Assessment of the iron load of serum ferritin was impaired by concomitant iron-containing proteins of similar size and shape, which could not be separated by SEC nor centrifugal ultra-filtration. In contrast, ferritin was the main iron-containing protein in cytosolic extracts of murine brain, which showed a total ferritin-bound iron content of (1.05 ± 0.12) µg g−1 (n= 10; U, k= 2). The relative expanded uncertainty achieved for a mass fraction of ca. 1 µg g−1 ferritin-bound iron was 11% (U rel, k = 2). The relative expanded uncertainty of the iron mass fraction of the (57Fe)ferritin spike was 5.7% and represented the major contributing factor to the overall uncertainty. Statistical tests suggested no significant difference in ferritin-bound iron content between mouse brain hemispheres. The presented analytical tool provides low limits of quantification (2.2 ng g−1) and uncertainties (11%, U rel, k = 2), thus enables the quantification of ferritin-bound iron in murine brain extracts with high sensitivity and accuracy. Furthermore, this analytical workflow assures comparability of measurement results across research laboratories. This provides the basis for investigation of the iron loading of ferritin in brain tissue of healthy and Alzheimer's disease mouse models, which may help answering the question if iron regulation is impaired in Alzheimer's disease.

中文翻译：

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使用物种特异性同位素稀释质谱法对用于阿尔茨海默病研究的鼠类样本中铁蛋白结合铁进行定量

我们研究了物种特异性同位素稀释质谱 (IDMS)，用于量化小鼠血清和大脑中铁蛋白结合的铁。因此，使用尺寸排阻色谱电感耦合等离子体质谱法 (SEC-ICP-MS) 分析新鲜样品。同位素标记的 (57Fe) 铁蛋白用作校准物，用于量化小鼠样品中铁蛋白结合的铁。血清铁蛋白铁负荷的评估受到伴随的类似大小和形状的含铁蛋白质的影响，这些蛋白质无法通过 SEC 或离心超滤分离。相比之下，铁蛋白是鼠脑细胞溶质提取物中的主要含铁蛋白质，其总铁蛋白结合铁含量为 (1.05 ± 0.12) µg g-1（n=10；U，k=2）。质量分数为约 1 的相对扩展不确定度。1 µg g-1 铁蛋白结合铁为 11% (U rel, k = 2)。(57Fe) 铁蛋白尖峰的铁质量分数的相对扩展不确定度为 5.7%，是总体不确定度的主要影响因素。统计测试表明，小鼠大脑半球之间的铁蛋白结合铁含量没有显着差异。所提出的分析工具提供了低定量限 (2.2 ng g-1) 和不确定性 (11%, U rel, k = 2)，从而能够以高灵敏度和准确度对鼠脑提取物中的铁蛋白结合铁进行定量。此外，这种分析工作流程确保了研究实验室之间测量结果的可比性。