微生物鉴鉴定系统-质谱法

Fontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings.AXIMA–iDPlus:质谱法快速鉴定微生物AssuranceConfidencePerformanceResonanceMALDITOF产品系列•Linearonly•MALDI-TOF•Linearandreflectron•MALDI-TOF•PSDcapability•Linearandreflectron•MALDI-TOF/TOF•PSD&CIDcapability•Uniqueinstrument•MALDI-iontrap-TOF•reflectrononly•MSncapability•HighresolutionandhighmassaccuracySampletargetIonopticsLineardetectorLaserMALDI-TOFMALDI:线性工作原理1.Mixmatrix&sampleontarget2.Firelaseratmatrix/sample3.Ionizesample(solidtogasphase)4.Accelerate/extractions5.Separateionsinanalyzer6.RecordspectrumatdetectorTimeofflight(TOF)massanalyzer(fieldfree)02040608010010000200003000040000m/z+vemode[M+H]+ionsMALDI基质辅助激光解析附原理:Streptococcuspneumoniae:m/z%Intensity95176865L327985L296638839458725263L346265L3065054419L369071S1880617703L3559516750L2810089L2710618S1910403S15475712310L7/L129357L31728611019S630005000700090001100013000880611451S105651L3310783L23masssignalsmatchingtoribosomalproteins(consideringpossiblemethioninecleavageand/ormethylation)aremarkedinred~50%ofpeaksunambiguouslyrepresentribosomalproteinsproteinmassesobtainedfromtheSwiss-Prot/TrEMBLsequencedatabaseFontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings微生物鉴定方法感官视觉,嗅觉,味觉免疫单或多克隆抗体分子PCR……生化代谢能力和耐抗生素耗时气质法（GC-MS）脂肪酸，小分子糖MALDI-TOFMSFontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings30-Oct-196微生物鉴定新方法（MALDITOF）•比现有方法鉴定更多生物多样性的病原体•分析时间仅1-2minutesFontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings现代微生物分类学:蛋白组学&基因组学ProteinBacterialCellDNAGenomeProteomeDNAFingerprint30005000700090001100013000Mass(m/z)2117.30102030405060708090100%IntensityVoyagerSpec#1=AdvBC(32,0.5,1.0)=NF0.7[BP=4423.9,2117]4424.29476.24861.08823.55897.86271.28054.36629.07424.912271.44738.66984.68383.86786.76533.39075.23132.36137.37209.88903.38610.610110.84765.410975.24026.94535.63311.810493.93491.49611.5MassFingerprintSpeciesDefinition&DeterminationMALDI-TOFMS88不同细菌代表性指纹图谱不同各菌种代表性峰型分布存在显著差异，有助于鉴定各个菌种！4000m/z80004000m/z8000BurkholderiacepaciaRaoultellaornithinolyticaStaphylococcusaureusPantoeaagglomeransAcinetobacterlwoffiEscherichiacoli99肠杆菌（Enterobacteriaceae）MS图谱示:科,属,种特异性峰值部分峰值是多数菌种共有的（如m/z4365:核糖体蛋白L36)峰值有科,属,种或亚种-特异性种间特异性代表峰值用来区分不同菌种30005000700090001100013000m/zEscherichiacoliHafniaalveiLeclerciaadecarboxylataKlebsiellapneumoniaeProteusmirabilisEnterobacteraerogenesFontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings核糖体蛋白不受生长条件影响MassspectraofProteusmirabilisDSM4479atdifferentincubationtimesNomajorqualitativechangesinpeakpatternsdistinguishingfamily,genusandspeciesDecreaseofsignalintensityovertimePresenceandabsenceofpeaksIdentificationwith99%confidenceFontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings样品SARAMIS工作流程转移细胞到MALDI靶板导入数据至SARAMIS4000600080001000012000m/z1[c].2I45263963877748176997060097545498240868962101657453125284478114226260LIMS鉴定/比较分析基质Fontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings微生物鉴定流程生长在固体培养基上的菌落全细胞MALDI-TOF谱图数据库中参考谱图输出可信鉴定结果找到匹配度最可信的谱图13鉴定结果界面样品名称鉴定结果置信度MS数据日期只需数秒，全自动添加新采集数据至鉴定结果界面！科属种型数据数量不同颜色代表不同置信水平深绿:99.9%浅绿:90-99.9%黄色:85-90%白色:70-85%低于70%不显示置信值红色:提醒复杂的结果匹配2种或以上菌种，置信度99%:鉴定混合菌Fontsize:Sizescanbeadjusteddependingonthetextlength.CharactersshouldnotbeoverlappedonglobalstringsSARAMIS鉴定混合菌种AmassspectrumofamixedsamplecontainingPseudomonasaeruginosaandAchromobacterxylosoxidansShowspeaksspecifictobothspecies300040005000600070008000900010000110001200013000Mass/Charge7291.64331.87319.65237.04973.76320.26692.27575.06618.34433.77097.78562.09086.06345.86045.05735.69402.710029.95209.06674.27487.87201.05014.67763.48880.73161.14700.96306.03787.23645.04543.14353.03344.55266.48745.86714.75676.09588.68403.99959.910123.29427.912565.810534.69215.88584.69724.211401.7单核增生性李斯特菌Listeriamonocytogenes(Lm)兼性厌氧的革兰氏阳性杆菌，自然界分布广泛，是食源性李斯特病（人畜共患病）的致病原因，低温仍具有增殖能力对老年人，婴儿，孕妇和免疫缺陷者的致死率是20-30%该菌经常存在于速食食品中Ex.About2,500cases/yearinUSA食品安全TokyoMarineUniversity,ProfKimuraandDr.TakahashiEatablewithoutheatedandcockedorcockedfood,ex.,ham,dailyproduct,salad,smokedfoodetc•现有检测方法需要2步骤和选择培养基，耗时6天Yakult(养乐多)乳酸菌生产质量控制品种改良Performance+SARAMISClassifiedasgenus=species=strain“Thickness”ofdatabaseisimportant.From2011Europeansocietyofclinicalmicrobiologyandinfectionsdisease菌种鉴定准确度Fontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeoverlappedonglobalstrings30-Oct-1918SARAMIS：SpectralARchiveAndMicrobialIdentificationSystem•数据库含有超级谱图（SuperSpectra）和参考谱图（ReferenceSpectra）•使用者可以创建和储存超级谱图或参考谱图•软件包可实现谱图的存储，分析和比较•使用者可以做聚类分析，创建系统树图用于菌株分型Fontsize:Sizescanbeadjusteddependingonthetextlength.CharactersshouldnotbeoverlappedonglobalstringsSARAMIS菌种特异峰型•合成谱图包含所有在个体谱图中出现频率大于阀值的所有峰(如：在各谱图中出现70%的)•在超级谱图中，所有峰值根据他们对所代表的菌种特异性而赋予权重值•具有科、属特异性的峰值权重低，这些峰值不支持菌种鉴定•具有菌种特异性的峰值权重则会加重Fontsize:Sizescanbeadjusteddependingonthetextlength.Charactersshouldnotbeov