液相串联质谱在遗传代谢病高危新生儿检测中运用研究

目的研究液相串联质谱在遗传代谢病高危新生儿检测中的意义。方法留取560例临床疑似遗传代谢病新生儿的干血滤纸片,经含氨基酸、酰基肉碱内标的甲醇萃取,盐酸正丁醇衍生后,进行液相串联质谱分析。结果560例高危新生儿中检测出患儿48例,阳性率8．6％（48／560）,包括脂肪酸代谢病11例（22．9％）;氨基酸代谢病15例（31．3％）;有机酸代谢病22例（45．8％）。结论液相串联质谱可以通过检测血滤纸片中的不同酰基肉碱浓度,一次分析快速检测30多种遗传代谢病,对于遗传代谢病高危新生儿的临床诊断具有重要的价值。     

6137例临床患儿遗传代谢病串联质谱及气相色谱质谱数据分析

目的了解本实验室2013年度串联质谱及气相色谱诊断氨基酸、有机酸、脂肪酸氧化代谢性疾病数据分析,深入对遗传代谢病诊断的认识。方法汇总整理2013年1~12月份的6137例(男3678例,女2459例)临床患儿的串联质谱(LC-MS/MS)及气相色谱(GC-MS)检测数据及临床诊断进行分析。结果在6137例临床患儿共检测出氨基酸代谢病(AAD)、有机酸代谢紊乱(OAD)和脂肪酸氧化缺陷病(FAOD)共144例检出率2.3%(男84例,女60例)。其中氨基酸代谢病占43.8%(63/144例),有机酸代谢紊乱占48.0%(69/144例),脂肪酸氧化缺陷病8.4%(12/144例),检测出较多代谢异常的为甲基丙二酸血症(MMA)56例、高苯丙氨酸血症32例、戊二酸血症Ⅰ型9例、枫糖尿病及肉碱/酰基肉碱移位酶缺乏症(CACT)各6例。结论临床患儿遗传代谢病并不少见,科室以新生儿科、神经科、康复科、PICU最多见,地区以山东、江苏、河北、安徽省较多;其中甲基丙二酸血症、高苯丙氨酸血症、戊二酸血症Ⅰ型最多见。儿科医师对临床患儿应重视串联质谱(LC-MS/MS)及气相色谱(GC-MS)技术检测遗传代谢病,以达到早查、早治疗,降低临床患儿不可逆伤害的目的。     

串联质谱技术在脂肪酸氧化代谢病诊断中的应用研究

目的探讨利用串联质谱技术检测干血滤纸片中酰基肉碱水平,诊断脂肪酸氧化代谢病。方法对象为2941例临床遗传性代谢病高危儿童,利用串联质谱技术检测患儿干血滤纸片中酰基肉碱水平,结合临床资料和常规生化结果,进行脂肪酸氧化代谢病诊断。结果诊断了14例脂肪酸氧化代谢病(0.5%),其中肉碱棕榈酰转移酶Ⅰ缺乏症1例,肉碱棕榈酰转移酶Ⅱ缺乏症1例,短链酰基辅酶A脱氢酶缺乏症1例,中链酰基辅酶A脱氢酶缺乏症7例,极长链酰基辅酶A脱氢酶缺乏症2例,多种酰基辅酶A脱氢酶缺乏症2例。结论通过串联质谱技术检测干血滤纸片中酰基肉碱水平,可对部分脂肪酸氧化代谢病进行诊断。     

廊坊地区41062例新生儿遗传代谢病串联质谱筛查结果分析

目的 对廊坊地区41062例新生儿35种遗传代谢病串联质谱筛查情况进行总结分析。方法 对2020年1月至2020年12月廊坊地区出生的新生儿足跟血进行氨基酸代谢异常、有机酸代谢异常、脂肪酸代谢异常等35种遗传代谢病串联质谱检测,总结目前廊坊地区新生儿串联质谱筛查工作现状,比较廊坊地区与国内外其他地区遗传代谢病发病率及种类分布。结果 41062例新生儿中,共筛查出患病儿童18例,总检出率为1/2281,其中氨基酸代谢障碍性疾病9例（50%）,脂肪酸代谢障碍性疾病5例（27.78%）,有机酸代谢障碍性疾病4例（22.22%）。结论 串联质谱检测技术可以高效地对遗传代谢病进行筛查,做到早发现早诊断早治疗,是出生缺陷防控的重要措施。     

新生儿遗传代谢病串联质谱筛查带教思考

通过教学实践,使实习生和进修生熟练掌握新生儿遗传代谢病串联质谱（MS/MS）筛查的实验原理及仪器使用方法,了解应用MS/MS技术开展新生儿疾病筛查中常见氨基酸或肉碱异常及其临床意义,为新生儿遗传代谢病串联质谱筛查的大规模开展奠定坚实的基础。     

液相色谱-串联质谱法筛查漏诊的4例甲基丙二酸血症

目的 对现行液相色谱-串联质谱(liquid chromatography-tandem mass spectrometry,LC-MS/MS)法筛查的甲基丙二酸血症患者的临漏诊床、生化及基因特点进行研究,以期改进诊断技术.方法 本研究纳入既往新生儿筛查或高危筛查阴性的4例甲基丙二酸血症患者,采用其他生化检测及基因分析明确诊断.应用LC-MS/MS技术对干血斑总同型半胱氨酸补充检测.结果 4例患者于2个月至4岁发病,主要表现为重度贫血、脑积水、发育落后及皮肤色素沉着.例1和例2血液丙酰肉碱增高,尿液甲基丙二酸增高,基因分析证实为cblC型.例3血清总同型半胱氨酸增高,基因检测证实为cblG型.例4血清总同型半胱氨酸和尿液甲基丙二酸增高,基因分析证实为cblJ型.LC-MS/MS检测发现,4例患者干血斑总同型半胱氨酸均显著增高.结论 本研究报道了4例LC-MS/MS漏筛的甲基丙二酸血症,确诊了我国首例cblG和cblJ.LC-MS/MS技术可实现干血斑总同型半胱氨酸检测,提高诊断效率,减少漏诊.     

21 024例新生儿遗传代谢病串联质谱结果回顾分析

目的探讨串联质谱技术在新生儿遗传代谢病筛查和诊断中的作用和意义,统计并了解我市新生儿遗传代谢病发病率。方法收集2016年3月～2018年12月余姚市出生的新生儿足跟血干血滤纸片,通过串联质谱技术检测样本中氨基酸及酰基肉碱等指标,以筛查氨基酸、有机酸和脂肪酸代谢病共26种遗传代谢病,可疑阳性者进一步确诊。结果共筛查21 024例新生儿,可疑阳性769例,确诊8例,包括苯丙酮尿症4例,甲基丙二酸血症1例,瓜氨酸血症Ⅰ型1例,原发性肉碱缺乏症2例。结论应用串联质谱技术极大地推动了新生儿遗传代谢病筛查的发展,实现了新生儿遗传代谢病早发现早治疗的目的,从而避免患儿发生不可逆性的生长及智能发育障碍,提高人口素质。     

临沂地区新生儿遗传代谢病串联质谱结果分析

目的探讨串联质谱技术在新生儿遗传代谢病筛查和诊断中的作用和意义,统计并了解我市遗传代谢病发病率。方法收集2014年7月～2017年9月临沂市出生的新生儿足跟血干血滤纸片,通过串联质谱技术检测样本中氨基酸及酰基肉碱等指标,以筛查氨基酸、有机酸和脂肪酸代谢病共48种遗传代谢病,可疑阳性者进一步确诊。结果共筛查83 570例新生儿,可疑阳性2002例,确诊16例,包括氨基酸代谢病1例(高甲硫氨酸血症),有机酸代谢病6例(甲基丙二酸血症5例,戊二酸血症Ⅰ型1例),脂肪酸代谢病9例(原发性肉碱缺乏症7例、短链酰基辅酶a脱氢酶缺乏症2例)。结论应用串联质谱技术在全市范围内筛查并确诊16例病例,实现了新生儿遗传代谢病早发现早治疗的目的,有效保障儿童健康,降低出生缺陷,提高出生人口素质。     

两种非衍生化串联质谱检测试剂在新生儿遗传代谢病筛查中的应用比较

目的比较两种非衍生化串联质谱检测试剂(NeoBase? 1和NeoBase? 2)在新生儿遗传代谢病筛查中的应用情况。方法选择2020年6月至12月在西北妇女儿童医院进行遗传代谢性疾病筛查的表型正常新生儿滤纸干血片标本, 共计7295份。同时应用NeoBase? 1和NeoBase? 2试剂检测滤纸干血斑样本, 了解正常新生儿11种氨基酸和12种酰基肉碱水平分布, 并制定参考区间;采用配对Wilcoxon检验比较11种氨基酸和12种酰基肉碱水平之间差异;同时检测经临床确诊的23例6种遗传代谢病患儿的标本, 验证两种试剂对疾病判读的一致性。结果正常新生儿的11种氨基酸和12种肉碱水平经K-S单标本正态分布检验, 呈偏态分布(均P<0.001), 两种试剂检测的所有指标水平差异均有统计学意义(均P<0.001);以0.5～99.5百分位数确定两种试剂检测各种氨基酸和酰基肉碱的正常参考范围, 两种试剂确定的参考范围有重叠, 经临床确诊样本验证, NeoBase? 2与NeoBase? 1均检测到相应异常指标的改变, 两种试剂均能检测出疾病患儿, 检出率均为100%。结论 NeoB...     

246例癫痫患儿遗传代谢性疾病质谱技术筛查

目的通过对癫痫患儿遗传代谢病质谱技术筛查,提高遗传代谢病所致儿童癫痫的认识,为临床诊治提供理论依据。方法选取癫痫患儿246例纳入本研究,应用气象色谱-质谱(GC-MS)和串联质谱(MS/MS)技术对入选的癫痫患儿血和尿氨基酸、有机酸、脂肪酸进行筛查,同时行血、尿常规、肝肾功能、心肌酶、血糖、血气分析及头颅MRI、EEG等检查,部分病例行血氨、乳酸、丙酮酸、血同型半胱氨酸等检测。结果筛查的246例癫痫患儿中,根据血尿筛查的典型改变结合临床表现及其它检查最后确诊遗传代谢病25例,其中甲基丙二酸血症9例(伴同型半胱氨酸血症2例),酮性双羧酸尿症3例,戊二酸血症、丙酸血症、瓜氨酸血症Ⅱ型、肉碱缺乏症及多羧酶缺乏症各2例,尿素循环障碍、4羟基丁酸尿症及苯丙酮尿症各1例。结论遗传代谢病是癫痫儿童病因之一,尤其伴有智力运动障碍、急慢性脑病及难治性的癫痫的婴幼儿尽早行先天性遗传代谢筛查,以助早期诊断,合理治疗。     

Pilot study of gas chromatographic-mass spectrometric screening of newborn urine for inborn errors of metabolism after treatment with urease.

Gas chromatographic-mass spectrometric (GC-MS) techniques for urinary organic acid profiling have been applied to high-risk screening for a wide range of diseases, mainly for inborn errors of metabolism (IEM), rather than to low-risk screening or mass screening. Using a simplified procedure with urease-pretreatment and the GC-MS technique, which allows simultaneous determination of organic acids, amino acids, sugars and sugar acids, we performed a pilot study of the application of this procedure to neonatal urine screening for 22 IEM. Out of 16,246 newborns screened, 11 cases of metabolic disorders were chemically diagnosed: two each of methylmalonic aciduria and glyceroluria, four of cystinuria, and one each of Hartnup disease, citrullinemia and alpha-aminoadipic aciduria/alpha-ketoadipic aciduria. The incidence of IEM was thus one per 1477, which was higher than the one per 3000 obtained in the USA in a study targeting amino acids and acylcarnitines in newborn blood spots by tandem mass spectrometry. Also, 227 cases were found to have transient metabolic abnormalities: 108 cases with neonatal tyrosinuria, 99 cases with neonatal galactosuria, and 20 cases with other transient metabolic disorders. Two hundred and thirty-eight cases out of 16,246 neonates (approximately 1/68) were thus diagnosed using this procedure as having either persistent or transient metabolic abnormalities.     

Application of tandem mass spectrometry to neonatal screening of inherited metabolic diseases: focus on present developments

The recent evolution of tandem mass spectrometry allows to diagnose more than twenty inherited metabolic diseases within a single blood spot. Nowadays, it is technically possible to screen newborns for most of fatty acid oxidation, organic acid and amino acid disorders. An important number of prospective pilot studies, using tandem mass spectrometry, have been done worldwide. However, several technical, economical, medical and ethical problems are raised by these applications. This review is intended to focus on this technology and to resume results from the main international studies.     

基于代谢组学评价血液生物标志物对中枢神经系统肿瘤的诊断价值

目的 分析中枢神经系统肿瘤代谢组学图谱,评价小分子代谢物作为中枢神经系统肿瘤生物标志物的诊断价值.方法 本研究纳入126例中枢神经系统肿瘤患者和86例健康对照人群.通过液相色谱质谱联用技术(LC-MS/MS)检测血液中氨基酸和酰基肉碱,分析中枢神经系统肿瘤代谢组学图谱.用ROC曲线对小分子代谢物进行诊断效能分析.结果 中枢神经系统肿瘤代谢谱与健康人群有明显区别,19个指标有显著差异.肿瘤患者血液C2水平显著低于健康人群.Arg,C8和C18:2对中枢神经系统原发性肿瘤具有辅助诊断价值,ROC曲线下面积分别为0.868、0.895和0.756,灵敏度分别为82.6%、88.0%和60.9%,特异性分别为77.5%、80.8%和78.3%.结论 血液中小分子代谢物,包括氨基酸和酰基肉碱可作为中枢神经系统肿瘤辅助诊断新的生物标志物.     

Recent developments in the investigation of inherited metabolic disorders using cultured human cells.

Thepurpose of this paper is to share experience with our systems and review recent "in vitro" methods using intact cells (fibroblasts, amniocytes) in which entire metabolic pathways can be probed for inherited metabolic defects reflected by elevations of intermediates determined by tandem mass spectrometry, HPLC, or gas chromatography-mass spectrometry. Currently, one can explore the integrity of mitochondrial fat oxidation, peroxisomal degradation of methyl-branched fatty acids (e.g., pristanate), and the mitochondrial degradation of the branched chain amino acids (leucine, valine, and isoleucine). For many of the diseases, the specific defect can be recognized from the acylcarnitine profile resulting from incubation of the intact cells with stable-isotope-labeled precursors to the particular pathway. This approach has also been successful in identifying new inherited metabolic disorders, biochemical correlation with clinical phenotypes of individual defects, and sequential oxidation of fatty acids by peroxisomal-mitochondrial interaction.     

Metabolite profiles of human immunodeficiency virus infected CD4+ T cells and macrophages using LC-MS/MS analysis

Human immunodeficiency virus type 1 (HIV-1) infects both activated CD4+ T cells and macrophages. We tested if liquid chromatography-tandem mass spectrometry (LC-MS/MS) technology can monitor metabolic alterations induced by HIV-1 in the infected cells. Here we monitored glucose uptake and conducted LC-MS/MS-based metabolomic analysis in HIV-1 infected primary human CD4+ T cells and a macrophage model system: differentiated U1 (HIV-1 producing) and differentiated U937 (control) cells. HIV-1 infected CD4+ T cells have higher glucose uptake and increases in several metabolite pool sizes, whereas HIV-1 producing macrophages had substantial reductions in glucose uptake and steady state glycolytic intermediates. This data suggests that the two HIV-1 target cell types exhibit very different metabolic outcomes during viral production. This study also validates the LC-MS/MS technology as an effective metabolomic approach to monitor various metabolic alterations made by HIV-1 infection.     

神经接头蛋白Dok6下游作用蛋白的鉴定及初步功能分析

目的 Dok6下游作用蛋白的鉴定和初步功能分析。方法利用anti-Dok6的抗体进行免疫共沉淀实验,然后通过Western blot,银染分析,质谱鉴定以及后续的生物信息学预测及分析其潜在的生理功能。结果本实验得到了26个可能与Dok6发生直接或间接相互作用的蛋白,这些蛋白分别参与了体内众多的生理过程。为了缩小研究范围,选取了与神经系统发育及细胞内信号传导相关的9个蛋白作为研究对象进行后续的功能研究。结论通过质谱鉴定以及生物信息学综合分析得到的潜在Dok6下游蛋白将给研究Dok6的生理功能及阐述其相关分子机制提供重要的依据。     

Evaluation of urinary acylglycines by electrospray tandem mass spectrometry in mitochondrial energy metabolism defects and organic acidurias

We analyzed the urinary acylglycine excretion in 26 patients with mitochondrial energy metabolism disorders and in 55 patients with organic acidurias by electrospray tandem mass spectrometry (ESI-MS/MS), monitoring precursor ions of m/z 90. Urinary concentrations of the different acylglycines were quantified using deuterated internal standards. Normal values for the most important acylglycines were established. In MCAD and MAD (neonatal form) deficiencies, typical excretion patterns of urinary acylglycines were found in all the samples. In isovaleric aciduria, propionic aciduria, and 3-methylcrotonylglycinuria typical glycine conjugates were always found. Methylmalonic aciduria (mutase deficiency), multiple carboxylase deficiency, and 3-hydroxy-3-methylglutaric aciduria revealed pathological acylglycine profiles, even if not specific for the disease. In all these diseases acylglycine excretion seems to be less influenced by the clinical status than organic acid excretion. This method is a useful diagnostic tool for these metabolic disorders, complementary to organic acids and acylcarnitine profiles.     

Simultaneous Identification of Multiple β-Lactamases in Acinetobacter baumannii in Relation to Carbapenem and Ceftazidime Resistance,Using Liquid Chromatography-Tandem Mass Spectrometry

Shotgun proteomics using liquid chromatography-tandem mass spectrometry (LC-MS/MS) was applied to detect β-lactamases in clinical Acinetobacter baumannii isolates. The correlation of the detection of β-lactamase proteins (rather than PCR detection of the corresponding genes) with the resistance phenotypes demonstrated an added value for LC-MS/MS in antimicrobial susceptibility testing.     

Chemical markers for bacteria in extraterrestrial samples

Interplanetary missions to collect pristine Martian surface samples for analysis of organic molecules, and to search for evidence of life, are in the planning phases. The only extraterrestrial samples currently on Earth are lunar dust and rocks, brought back by the Apollo (U.S.) and Luna (Soviet Union) missions to the moon, and meteorites. Meteorites are contaminated when they pass through the Earth's atmosphere, and during environmental exposure on Earth. Lunar fines have been stored on Earth for over 30 years under conditions designed to avoid chemical but not microbiological contamination. It has been extremely difficult to draw firm conclusions about the origin of chemicals (including amino acids) in extraterrestrial samples. Of particular concern has been the possibility of bacterial contamination. Recent work using state-of-the-art gas chromatography tandem mass spectrometry (GC-MS/MS) has dramatically lowered the chemical background, allowing a clear demonstration that lunar fines are remarkably different from terrestrial dust in that they generally lack certain chemical markers (muramic acid and 3-hydroxy fatty acids) characteristic of Earth's bacteria. Thus, lunar dust might be used as a negative control, in conjunction with GC-MS/MS analyses, in future analytical studies of lunar dust and meteorites. Such analyses may also be important in studies designed to search for the presence of life on Mars.