Zinellu Angelo, Carru Ciriaco, Galistu Franca, Usai Maria Franca, Pes Giovanni Mario, Baggio Giovannella, Federici Giorgio, Deiana Luca
Chair of Clinical Biochemistry, University of Sassari, Viale San Pietro 43/B, I-07100 Sassari, Italy.
Electrophoresis. 2003 Aug;24(16):2796-804. doi: 10.1002/elps.200305570.
We describe an ultrarapid capillary electrophoresis with laser-induced fluorescence (CE-LIF) method for total plasma thiols measurement. Reduced thiols by 10% tri-n-butylphosphine (TBP) were derivatized in 10 min at room temperature with 5-iodoacetamidofluorescein (5-IAF) as fluorescent reagent. We show that CE-LIF allows a baseline separation of total plasma cysteinylglycine, homocysteine, cysteine, and glutathione in less than 5 min when N-methyl-D-glucamine in run buffer was added. CE was compared with high-performance liquid chromatography (HPLC) with fluorescence detection. The Bland-Altman test and Passing-Bablok regression demonstrates that the results obtained by CE-LIF and by HPLC are highly comparable. The simplified procedure of sample preparation, the short incubation and fast separation times, the high specificity, sensitivity and reproducibility, and the lower cost of analysis suggest that our proposed method can be considered valuable for the automation analysis in a routine laboratory.
我们描述了一种用于测量血浆总硫醇的超快速毛细管电泳结合激光诱导荧光（CE-LIF）方法。用10%三正丁基膦（TBP）还原的硫醇在室温下与5-碘乙酰氨基荧光素（5-IAF）作为荧光试剂反应10分钟进行衍生化。我们表明，当在运行缓冲液中添加N-甲基-D-葡糖胺时，CE-LIF能够在不到5分钟的时间内对血浆中的总半胱氨酰甘氨酸、同型半胱氨酸、半胱氨酸和谷胱甘肽进行基线分离。将CE与具有荧光检测的高效液相色谱（HPLC）进行了比较。Bland-Altman检验和Passing-Bablok回归表明，CE-LIF和HPLC获得的结果具有高度可比性。简化的样品制备程序、较短的孵育和快速分离时间、高特异性、灵敏度和重现性以及较低的分析成本表明，我们提出的方法对于常规实验室的自动化分析具有重要价值。
