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目的分析甲胎蛋白、瘦素及肌钙蛋白Ⅰ等重要生物标志物的绝对定量方式。方法采用高效液相色谱-同位素稀释串联质谱法对人源甲胎蛋白、人源瘦素以及重组肌钙蛋白予以测量,并对比分析其检测方法的准确性。数据比较采用t检验,P<0.05为差异有统计学意义。结果对4份平行样品予以酶解,并将酶解后的产物予以高效液相色谱-同位素稀释串联质谱检测,计算出肽段FK和NR浓度,最终AFP结果为5.45μmol/g,与水解法检测的结果相比,显示其相对偏差为6.8%。二者检测结果的一致性检验显示,差异无统计学意义(P>0.05)。人源瘦素含量为0.59 g/g,与水解法测出的0.63 g/g相比,相对标准偏差为5.21%~6.07%,处于误差允许范围内(t=0.89,P>0.05);肌钙蛋白Ⅰ的质量分数为0.24 g/g,与水解法测定的0.21 g/g比较,可知其相对偏差处于4.75%~8.43%,处于误差允许范围内(t=0.53,P>0.05)。结论高效液相色谱—同位素稀释串联质谱法检测人源甲胎蛋白、人源瘦素及重组肌钙蛋白Ⅰ的准确性高,灵敏度好,且可重复进行,具备推广应用价值。
Objective To analyze the absolute quantification of important biomarkers such as alpha-fetoprotein, leptin and troponin I. Methods Human alpha-fetoprotein, human leptin and recombinant troponin were measured by high performance liquid chromatography-isotope dilution tandem mass spectrometry. The accuracy of the detection method was compared. Data were compared using t test, P <0.05 for the difference was statistically significant. Results The four parallel samples were digested and the products of enzymatic hydrolysis were detected by high performance liquid chromatography-isotope dilution-tandem mass spectrometry. The FK and NR concentrations of the peptides were calculated. The final AFP value was 5.45μmol / g, The test results showed a relative deviation of 6.8%. Consistency test results between the two showed no significant difference (P> 0.05). The relative standard deviation was 5.21% ~ 6.07%, which was within the allowable range of error (t = 0.89, P> 0.05). The leptin content of human was 0.59 g / g. Compared with 0.63 g / Calcium protein I mass fraction was 0.24 g / g, compared with the hydrolysis of 0.21 g / g, we can see that the relative deviation of 4.75% ~ 8.43%, within the error allowed range (t = 0.53, P> 0.05). Conclusion High-performance liquid chromatography-isotope dilution-tandem mass spectrometry has the advantages of high accuracy, good sensitivity and reproducibility for the detection of human alpha-fetoprotein, human leptin and recombinant troponin I, and has the value of popularization and application.