摘要目的 建立同位素稀释液相色谱串联质谱法(ID-LC/MS/MS)测定血清甲状腺素的候选参考方法.方法 实验性基础研究.重量法准确地称取血清或校准溶液,稀释器加入13C6-甲状腺素,混合,除去蛋白后经过阳离子交换固相萃取柱,收集洗脱液氮气吹干,复溶后用LC/MS/MS测定甲状腺素和内标.用7种混合血清评价方法的精密度;以其中一种血清为样本,分别向样本中加入原样本浓度50％、100％和200％的标准物质评价方法的加样回收率;分别以信噪比(S/N) ＞3和(S/N)＞10作为标准,评价方法的检测限和定量限.分析德国临床化学会参考物质CRM 21201和CRM21202,评价方法的正确度.结果 血清甲状腺素测定的批内CV和总CV分别为0.64％ (0.35％ ～0.89％)和0.83％(0.57％ ～1.37％).加样回收率为99.6％ ～ 100.7％.检测限和定量限分别为0.12和0.41nmoL/L.分析CRM 21201和CRM 21202,测定结果与认定值的平均偏差为-0.30％(范围-0.13％～0.73％).结论 建立了ID-LC/MS/MS测定血清甲状腺素的方法,方法取样量少,测定结果正确、精密,可望作为血清甲状腺素测定的参考方法.

abstractsObjective An isotope dilution liquid chromatography tandem mass spectrometric (ID-LC/MS/MS) method was developed for serum total thyroxine.Methods Serum samples were sampled by weight and the 13C6-thyroxine internal standards were added volumetrically using automated dilutors,followed by equilibration,protein precipitation,and cation exchang solid-phase extractions (SPE).After SPE,the eluates were evaporated to dryness under nitrogen and then the evaporated residues were reconstituted to prepare samples for liquid chromatography-mass spectrometry electrospray ionization (LC/MS-ESI) analysis.Seven kinds of serum pools were used to analyze the precision of the method.The analytical recovery,the limit of detection(signal to noise ratio,S/N ratio 3∶1) and the limit of quantification (S/N ratio 10∶1)for thyroxine were calculated.Also,in order to evaluate the accuracy of this method,the candidate reference materials from German Societies for Clinical Chemistry were analyzed.Results The within-run and total coefficients of variation were:0.64％ (0.35％-0.89 ％) and 0.83％ (0.57％-1.37 ％),respectively.The analytical recoveries ranged from 99.6％ to 100.7％.The limit of detection was 0.12 nmol/L and the limit of quantification was 0.41 nmol/L for thyroxine in human serum.The results of analyzing the reference material of CRM21201 and CRM21202 showed biases of-0.30％ (ranged from-0.13％ to 0.73％).Conclusions A highly precise and accurate method for measuring serum total thyroxine has been developed by ID-LC/MS/MS.The method is less sample volume demand and less time-consuming and may be used as a candidate reference method.