Measurement of thyroglobulin by liquid chromatography-tandem mass spectrometry in serum and plasma in the presence of antithyroglobulin autoantibodies.
Measurement of thyroglobulin by liquid chromatography-tandem mass spectrometry in serum and plasma in the presence of antithyroglobulin autoantibodies.
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DOI：
10.1373/clinchem.2012.195594
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发表时间：
2013-06
影响因子：
9.3
通讯作者：
Meikle AW
中科院分区：
文献类型：
--
作者：
Kushnir MM;Rockwood AL;Roberts WL;Abraham D;Hoofnagle AN;Meikle AW
血清甲状腺球蛋白（Tg）的测量用于监测分化型甲状腺癌（TC）治疗后的患者。使用Tg作为许多患者TC复发的生物标志物的困难源于内源性抗Tg自身抗体（Tg-AAb）的存在，其可干扰免疫测定（IA）并导致假阴性结果。使用兔多克隆抗Tg抗血清和蛋白沉淀从血清样品中富集Tg。不相关的蛋白质在此过程中部分耗尽。富集的蛋白质，然后变性，还原，并消化后，用胰蛋白酶的翅膀内标肽。通过免疫亲和提取纯化Tg特异性胰蛋白酶肽，并通过2D-LC-MS/MS进行分析。每个样品的仪器循环时间为6.5 min。定量下限为0.5 ng/mL（0.76 fmol/mL二聚体）。血清样本在5天内重复测定3次的总不精密度小于10%。与使用不含Tg-AAb的血清样品（n=73）的市售IA比较，显示德明回归，IA= 1.00*LC-MS/MS-2.35，r=0.982，Sy，x=9.52。在一组使用IA检测Tg为阴性的Tg-AAb阳性样品（n=71）中，通过LC-MS/MS法测定的浓度在23%的样品中等于或高于0.5（中位数1.2，范围0.7-11 ng/mL）。该方法具有用于临床诊断应用的可接受的性能特征。在浓度低于2 ng/mL（采用LC-MS/MS测定）的Tg-AAb阳性样品中观察到方法之间最显著的差异。该方法中用于Tg的亲和辅助富集策略适用于具有内源性自身抗体的其他生物标志物。
Measurement of serum thyroglobulin (Tg) is used to monitor patients after treatment for differentiated thyroid carcinoma (TC). Difficulty in using Tg as a biomarker of the recurrence of TC in many patients stems from the presence of endogenous anti-Tg autoantibodies (Tg-AAb), which can interfere with immunoassays (IA) and cause false-negative results. Tg was enriched from serum samples using rabbit polyclonal anti-Tg antiserum and protein precipitation. Unrelated proteins were partially depleted in the process. Enriched proteins were then denatured, reduced, and digested with trypsin after the addition of a winged internal standard peptide. A Tg-specific tryptic peptide was purified by immunoaffinity extraction and analyzed by 2D-LC-MS/MS. Instrument cycle time was 6.5 min per sample. The lower limit of quantification was 0.5 ng/mL (0.76 fmol/mL of dimer). Total imprecision of triplicate measurements in serum samples over five days was less than 10%. Comparison with a commercial IA using serum samples free of Tg-AAb (n=73) showed Deming regression, IA= 1.00*LC-MS/MS-2.35, r=0.982, Sy,x=9.52. In a set of Tg-AAb positive samples tested negative for Tg using IA (n=71), concentrations determined by LC-MS/MS method were at or above 0.5 in 23% of samples (median 1.2, range 0.7–11 ng/mL). The method has acceptable performance characteristics for use in clinical diagnostic applications. The most substantial disagreement between the methods was observed in Tg-AAb positive samples with concentration below 2 ng/mL (determined with LC-MS/MS). The affinity assisted enrichment strategy used for Tg in this method is applicable to other biomarkers that have endogenous autoantibodies.
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影响因子：
9.3
作者：
Okosieme, OE;Evans, C;Lazarus, JH
通讯作者：
Lazarus, JH
影响因子：
4.4
作者：
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5.8
作者：
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Schalin-Jantti, Camilla
影响因子：
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作者：
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通讯作者：
Meikle, A. Wayne
影响因子：
1.7
作者：
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通讯作者：
Ain, KB