干滤纸血片制备因素对遗传代谢病串联质谱分析结果的影响  

Investigation of the Factors Affecting the Preparation of Dry Blood Spot on Tandem Mass Spectrometry Analysis Resμlts for Inherited Metabolic Disorders

作　　者：周启凤 曹闻琦 吴虹林 和秀梅 何玺玉 ZHOU Qifeng;CAO Wenqi;WU Honglin;HE Xiumei;HE Xiyu(College of Medicine of Chinese People,s Liberation Army,Beijing 100853,China;Fifth Medical Center,Chinese People,s Liberation Army General Hospital,Beijing 100853,China)

机构地区：[1]中国人民解放军医学院,北京100853 [2]中国人民解放军总医院第五医学中心,北京100853

出　　处：《标记免疫分析与临床》2025年第1期123-128,共6页Labeled Immunoassays and Clinical Medicine

基　　金：国家重点研发计划项目(编号:2022YFF0710304)。

摘　　要：目的探究新生儿遗传代谢病筛查实验室分析前环节中,干滤纸血片制备时可能影响串联质谱分析结果的因素。方法将所收集全血的红细胞压积调整为50%~55%,向其中富集瓜氨酸、甲硫氨酸、苯丙氨酸、游离肉碱、乙酰肉碱、丙酰肉碱工作液,用移液枪吸取后滴落至滤纸上,将干滤纸血片样本按血斑点样血量(10、25、35、50、75、100μL)分6组,测量各组血斑直径。采用LC-MS/MS方法分析25、35、50、75、100μL组样本内主要氨基酸(甲硫氨酸、苯丙氨酸、瓜氨酸)和酰基肉碱(游离肉碱、乙酰肉碱、丙酰肉碱)标志物浓度水平。比较50μL组血片样本不同部位(中央和边缘)的主要标志物浓度变化。结果直径均大于8mm的血斑样本组主要氨基酸、酰基肉碱浓度比较存在差异。25μL组样本内主要标志物浓度均显著小于50μL组、75μL组、100μL组(P均<0.05);50μL组和75μL组样本内各标志物水平相当,差异未见统计学意义;100μL组样本中主要标志物浓度均显著大于其余各组(P均<0.05)。在干血片边缘部位的主要标志物浓度水平高于中央部位,差异有统计学意义(P<0.05)。结论在氨基酸和酰基肉碱谱的实验室分析前环节中,干滤纸血片制备时不同血斑点样血量和取样部位会对串联质谱分析结果造成影响,在制备血片时,推荐点样血量为50~75μL,取样部位应选取在血斑边缘部位进行。Objective To explore factors that could affect the results of tandem mass spectrometry analysis in the preparation of dry blood filter paper in the pre-analysis of neonatal inherited metabolic diseases screening laboratory.Methods The hematocrit of the collected whole blood samples was adjusted to 50%-55%,and citrulline,methionine,phenylalanine,free carnitine,acetyl carnitine and propionylcarnitine were enriched into the blood.The blood sample was sucked up with a pipette and dripped onto the filter paper,and the blood samples of the dry filter paper were divided into 6 groups according to the blood spot blood volume(10,25,35,50,75,100μL).The diameter of the blood spots in each group was then measured.LC-MS/MS was used to measure the concentration levels of the main amino acids(methionine,phenylalanine,citrulline)and acylcarnitine(free carnitine,acetyl carnitine,propionylcarnitine)markers in the samples of 25μL,35μL,50μL,75μL and 100μL groups.The changes in the concentration of the main markers in different parts(e.g.central and marginal)of blood samples in the 50μL group were compared.Results There were differences in the concentrations of major amino acids and acylcarnitine among the blood spot samples with a diameter of more than 8 mm.The concentrations of the main markers in the 25μL group were significantly lower than those in the 50μL,75μL and 100μL groups(all P<0.05).The levels of each marker in the samples of the 50μL group and the 75μL group were similar,while the difference was not statistically significant.The concentrations of the main markers in the 100μL group were significantly higher than those in the other groups(all P<0.05).The concentration level of the main markers at the edge of the dried blood film was significantly higher than that at the central site(P<0.05).Conclusion In the pre-laboratory spectrometry analysis of amino acids and acylcarnitine,the blood volume and sampling site of different blood spots can affect the results of tandem mass spectrometry analysis during the pr

关 键 词：新生儿筛查 遗传代谢病 干血斑 氨基酸 酰基肉碱 

分 类 号：R392-33[医药卫生—免疫学]