自動血液分析儀---SYSMEXXE-2100COULTERGen.S病理檢驗部一般檢驗科簡介更多的分析項目C:WBC,RBC,HGB,HCT,MCV,MCH,MCHC,PLT-ORDW-SD,RDW-CV,PDW,MPV,PCT,P-LCRD:NEUT%,LYMPH%,MONO%,EO%,BASO%NEUT#,LYMPH#,MONO#,EO#,BASO#N:NRBC%,NRBC#R:RET%,RET#,HFR,MFR,LFR,IRF,PLT-I32個分析項目,分別歸為4類32分析項目選擇選擇不同的分析方式1.C2.C+D3.C+N4.C+D+N5.C+R6.C+D+R7.C+D+N+RC:CBCD:DCN:NRBCR:RETIC操作更方便使用薄膜式按鍵即可完成所有例行作業(yè)關機聯(lián)機求助多功能鍵Sampler上機輸入Manual上機輸入光標IPU檢體量(自動穿刺)(開蓋分析)全新的偵測管路及分析原理ParametersPrincipleWBC,BASONEUT&BASO,LYMPH,MONO,EONRBCRET,IRF,PLT-OFlowCytometryusingSemiconductorlaserRBC,HCT,MCV,PLT-IHydrodynamicfocusingHGBSLS-HbHPC&ImmatureCellRF/DCDetectorWBC/BasoDIFFNRBCRETRBC/PLTHGBIMI四種偵測原理,七個偵測管路染料染料的成份

Polymethinor/andOxazine染色標的

Polymethine:DNA+RNAOxazine:DNA主要分析項目原理介紹RBCHgBPLTWBCDIFFIMINRBCRETRBC分析流程BloodCELLPACKRBC/PLTDetectorBlockSRV2,000μL-Blood:4.0μL-Reagent:1,996μLRBCsamplechamberCELLSHEATHImpedance(DCMethod)ExternalElectrodeApertureInternalElectrode電位=阻抗x電流當電流固定時,阻抗與電位成正比血球占去相等體積的稀釋液空間造成阻抗的改變HydrodynamicFocusing原理前后高壓導流導正血球排列及方向流體動力聚焦Hb的偵測偵測原理:SLSMethod/Spectrophotometer偵測位置:HbChannel提供項目:HbSLS試藥反應SLS厭水端與球蛋白結(jié)合立體結(jié)構(gòu)的改變氧化:Fe2+-->Fe3+SLS親水端與Fe3+鍵結(jié)-->SLS-HbFe2+Fe3+Fe3+SLSSLS-Hb優(yōu)點1.SLS(SodiumLauryalSulfate)是一種Detergent&Surfactant。2.與標準氰化物法相關性高(R2=0.999)3.Cyanidefreereagent；無生物毒性。4.Formadehydefreereagent；不刺激呼吸系統(tǒng)。5.沒有Oxyhemoglobinmethod無法氧化Methemoglobin

的缺點。Impedance/HydrodynamicFocusing原理PLT-I的HistogramFCM/Laser/Fluorescence原理FlowCellCELLPACKCELLPACKSampleSemiconductorLaserLaserbeam:633nmFCM/Laser/Fluorescence原理側(cè)散亂光RNA,DNA的含量細胞內(nèi)顆粒,結(jié)構(gòu),復雜性細胞的大小前散亂光螢光激光光DichroicmirrorPLT-oScattergramForwardScatter(大小)Fluorescence(DNA,RNA)GiantPLTFragmentedRBCSmallPLTXE-2100利用RNA含量的差異,PLT含RNA,成熟RBC不含以光學原理區(qū)分RBC與PLT,使其成為不連續(xù)的分布WBC,DIFF偵測偵測原理:FCM/LaserScatterLight/Fluorescence-偵測前散亂光,側(cè)散亂光

-偵測DNA,RNA含量偵測位置:1.WBC/BasoChannel2.DIFFChannel提供項目:WBC#,Nut#&%,Lym#&%,Mono#&%Eo#&%,Ba#&%DIFF試藥反應-normalcellLymMonoNeut+BasoEo1.針對Eosinophil以Organicacid處理,加強SideScatterLight,以增加Eo偵測敏感度2.中性試劑,含兩種接口活性劑,能將RBC溶解,而白血球則只有輕微的皺縮,抗溶解RBC沒有RNA,

也沒有顆粒,而儀器忽略“FSc(大小)＂的偵測,

因此沒有紅血球破壞不全的現(xiàn)象,不會干擾分類反應前反應后SSc(內(nèi)容物)Flu(RNA)DIFFScattergram(DNA,RNA)(內(nèi)容物結(jié)構(gòu))DIFF試藥反應-abnormalcell反應前反應后ImmatureWBC含有較多的RNA成份,因此螢光反應較成熟的WBC強DIFFScattergram(abnpattern)SideScatter(Internalstructure)BandMetaMyeloProMyeloBlastAty-LymFluorescence(RNA,DNA)DIFFScattergaramNORMALABNORMALImmatureWBC偵測XE-2100提供兩種分析原理偵測ImmatureWBC

偵測原理1:FCM/LaserScatterLight/Fluorescence

以“RNA含量＂,“細胞內(nèi)結(jié)構(gòu)信息＂偵測偵測位置:DIFFChannel偵測原理2:RF/DC

以“細胞膜脂質(zhì)含量＂,“細胞核密度＂偵測偵測位置:IMIChannel提供項目:Blast?,IG?,LS?,Aty-Ly?,HPC#&%,IG#&%(HPC:HaematopoeticProgenitorCell,造血前驅(qū)細胞,相當于CD34+Cell

也就是PBSC,周邊血液干細胞)ImmatureWBC偵測原理-1SideScatter(Internalstructure)BandMetaMyeloProMyeloBlastAty-LymFluorescence(RNA,DNA)以“RNA含量＂,“細胞內(nèi)結(jié)構(gòu)信息＂偵測ImmatureWBC偵測原理-2RF:RadiofrequencyDC:DirectCurrent(Impedance)細胞的大小細胞核密度IMI試藥反應(反應前)IMI試藥反應反應后,成熟的WBC裸核化,ImmatureWBC形態(tài)輕微皺縮IMI試藥反應成熟白血球不成熟白血球IMIScattergram細胞在成熟的過程中最大的差異在于核密度的變化利用RF來偵測細胞核的密度可以很敏感的將Blast,IG,band等分開RFDCBandMyelocyteMetamyelocyteBlastPromyelocyte以“細胞膜脂質(zhì)含量＂,“細胞核密度＂偵測AbnormalCell分布BandMetaMyeloProMyeloBlastAty-LymFluorescence(RNA,DNA)SideScatter(Internalstructure)PLTClumpImmatureGranBlastHPCLeftshiftRF(細胞核的密度)DC(大小)ABA可偵測Lymphoblast/MyeloblastB可偵測MyeloblastA+B+MyeloblastA+B-LymphoblastXE-2100提供Blasts?以及L-Blast?兩種FlagIMIDIFFnormalLymQ-FlagQ-Flag為儀器對SuspactMessage“Possibility＂的Index可用在儀器與手工鏡檢比對時,調(diào)整敏感度畫面中數(shù)字為Q-FlagIndex,代表Possibility定量NRBC的偵測偵測原理:FCM/LaserScatterLight/Fluorescence-偵測前散亂光/側(cè)散亂光

-偵測DNA,RNA含量偵測位置:NRBCChannel提供項目:NRBC#,NRBC%(NRBC/100WBC)NRBC試藥反應反應前反應后特殊酸性試劑,含水楊酸成份,能加強RBC的溶解,而WBC則輕微的皺縮,同時作核酸的染色NRBC試藥反應WBCNRBCRBCHighLevelFluorescenceSize:BigLowLevelFluorescenceSize:SmallWeakFluorescenceSize:MicroLysing(WBCmembraneisweaklydamaged)StainingRNADNANRBCScattergram(DNA,RNA)(大小)“Achangeoccursinelectricalresistancebetweentwoelectrodeswhenanonconductiveparticlesuchasabloodcellpassesbetweenthem＂ -WallaceCoulter,1947TheCoulterPrinciple“ScienceServingHumanity.＂AcTAcT5PDHmXSTKSGenSLH755AcT5PDALCountingtheCellsCellsaredilutedinanelectricallyconductivefluid(0.085%saltwater).Electrodesareseparatedbyasmallholethroughwhichthecurrentflows.Asthefluidandcellsarepulledthroughthehole,thecelltemporarilyblockstheflowofcurrentandcreatescountablepulses.SizingtheCellsThelargerthecell,themoreelectricalcurrentitblocks.Theheightofthepulseisequaltothevolumeofthecell.TheCoultermethodcountsandsizescellsbydetectingandmeasuringchangesinelectricalresistancewhenaparticle(suchasacell)inaconductiveliquidgoesthroughasmallaperture.#ofPulseParticlecountSizeofPulseCellVolumeCBCAnalysis-ImpedancePrincipleSensingZoneRedBloodCellAredcellpassesthroughRBCapertureTheCoulterprincipleOscilloscopeOscilloscopeOscilloscopeSensingZoneNeutrophilAWhitecellpassesthroughWBCapertureTheCoulterprincipleOscilloscope20fl2flBaselineRBCofvarioussizesPlateletsApplyingThresholdstoseparatePLTfromRBCHydrodynamicFocusing檢體不會與Sheathfluid混合確定檢體通過偵測孔正中央防止回流HydrodynamicFocusingwithDiluentSheath8psiCellsApertureSensingZoneRBCspushedawaybydiluentOscilloscopescreenRBCPLT+-DiluentstreamSystemwithsweepflow

掃流系統(tǒng)SweepFlowPulsetobeeditedDiluentstreamPulseediting(脈沖修正)NoncentralcellflowHemoglobin(HGB)國際標準法:氰化高鐵血色素法原理溶解紅血球?qū)⒍r鐵氧化為三價鐵之高鐵血色素高鐵血色素與KCN結(jié)合為氧化高鐵血色素

540nm比色優(yōu)點專一性佳缺點

KCN具毒性且造成環(huán)境污染

光學比色法原理Cyanide-freereagentBeer＇sLaw測定血紅素稀釋液的吸光度變化吸光度波長為540nm血紅素復合物為吸收光線的主要成分MCV直接測定法將雙凹盤狀紅血球球狀化直接偵測紅血球通過時發(fā)生電阻波形之高度推算之體積避免deformedRBCMCV偏低較傳統(tǒng)計算法更能看出紅血球?qū)嶋H大小PackedCellVolume(PCV;Hematocrit)WintrobetubemethodMicrohematocritmethodAutomaticmethodRedcelldistributionwidth(RDW)反應紅血球大小分布及差異性PriceJohns曲線圖直徑測定法自動血球計算法Anisocytosis

RDW-CV=×100%MCV

SD血小板測定方法

人工計數(shù)法電子阻抗法光學測定法免疫血小板電子阻抗法

自動血球分析儀最常見的原理以Hydrodynamicfocusingflow,測單一方向大小

將特定大小范圍內(nèi)的物質(zhì)定為血小板白血球計數(shù)及分類測定需將紅血球溶解以阻抗法測數(shù)目分類使用ScatterLight偵測

(3-partdifferential)CoulterWBCHistogram

(3-partdifferential)LymMonoEosNeutBaso

WBC50100200300400fLLyEoNeRBCErythroLyseStabliLyseNearNativeFlowcyometryVCSTechnologyDirect5-partDifferentialCountTechnologyRecap最佳血球大小偵測方法偵測0~3