Background:

Myeloperoxidase(MPO)isaglycoproteinpresentintheazurophil(primary)granulesofmyeloidcells,whichappearsinthemyeloblaststageofmyeloidcelldifferentiation.MPOishemostcommonfunctionalproteinofmyeloidcellsandisinvolvedintheinflammatoryresponse.Ithelpstokillmicrobesbybreakingdownperoxideinthepresenceofhalideions,contributingtothebactericidalfunctionofgranulocytes.TheprimarytranslationproductofMPOundergoesglycosylationwithproductionofthe89kDaheme-freeapopro-MPOformfollowedbyincorporationofhemeandconversionintotheenzymaticallyactivepro-MPOform.Subsequently,pro-MPObecomestargetedtoazurophilgranuleswherefinalprocessingoccurstoproducematuredimericMPOconsistingofthe59-64kDaMPOα-chainandthe14kDaMPOβ-chain.Lactoferrin(LF)isaniron-bindingproteinwithbactericidalandbacteriostaticactivitywhichisstoredwithinthesecondarygranulesofgranulocytes.LFexpressionisrestrictedtothepost-mitoticmaturationcompartmentofthegranulocyticlineage,startingfromthemyelocytestage.NormalandmalignantmyeloblastsareLFnegative.ThecombinedstainingforMPOandLFallowsthedistinctionbetweenmatureandimmaturemyelomonocyticcells.TheMPO-C2/LFCOMBI-ICreagentpermitstheidentificationandenumerationofimmatureandmorematuremyelomonocyticcellpopulationsinnormalandmalignanthumanbloodandbonemarrowusingflowcytometry.Resultsmustbeputwithinthecontextofotherdiagnostictestsaswellastheclinicalhistoryofthepatientbyacertifiedprofessionalbeforefinalinterpretation.Analysesperformedwiththisantibodyshouldbeparalleledbypositiveandnegativecontrols.Ifunexpectedresultsareobtainedwhichcannotbeattributedtodifferencesinlaboratoryprocedures,pleasecontactus.

Product:

1mlofFITC-conjugatedantiMyeloperoxidase-C2(clone8E6)andPE-conjugatedantiLactoferrin(clone4C5)inPBSpH7.2,1%BSA,and0.05%NaN3,approximately50tests.

Applications:

PermeABIlizationandStainingProcedure-IncombinationwithourPermeabilizationKitFIX&PERM(Cat.No.GAS-002)intracellularMPO-C2andLFcanbeeasilystainedincellsUSPensions.-Foreachsampletobeanalyzedadd50µlofwholeblood,bonemarrowormononuclearcellsuspensionina5mltube-Add100µlofReagentA(FixationMedium,storedandusedatroomtemperature)-Incubatefor15minutesatroomtemperature-Add5mlphosphatebufferedsalineandcentrifugecellsfor5minutesat300g-Removesupernatantandaddtocellpellet100µlReagentB(PermeabilizationMedium)and20µloftheMPO-C2/LFCOMBI-ICmonoclonalantibodyconjugate-Vortexatlowspeedfor1-2seconds-Incubatefor15minutesatroomtemperature-Washcellswithphosphatebufferedsalineasdescribedabove-Removesupernatantandresuspendcellsinsheathfluidforimmediateanalysisorresuspendcellsin0.5ml1.0%formaldehydeandstorethemat2-8°Cinthedark.Analyzefixedcellswithin24hours

Specificity:

AntibodyMPO-C2(clone8E6)reactswithhumanmyeloperoxidase(MPO)expressedbynormalandmalignantmyelomonocyticcells.TheLFmAb(clone4C5)recognizeslactoferrinstoredwithinsecondarygranulesofpostmitoticgranulocyte-commitedcells.InthisCOMBI-ICReagentantibody8E6isconjugatedtoFITC,antibody4C5isconjugatedtoPhycoeythrin(PE).ThesensitivityofMPO-C2/LFmAbisdeterminedbystainingwell-definedbloodsamplesfromrepresentativedonorswithserial-foldmAbdilutionstoobtainatitrationcurvethatallowsrelatingthemAbconcentrationtothepercentageofstainedcellsandgeometricMFI(meanfluorescenceintensity).Forthispurpose,amAb-concentrationrangeisselectedtoincludeboththesaturationpoint(i.e.themAbdilutionexpectedtobindallepitopesonthetargetcell)andthedetectionthreshold(i.e.themAbdilutionexpectedtorepresenttheleastamountofmAbneededtodetectanidenticalpercentageofcells).Inpractice,50µlofleukocytescontaining10^7cells/mlarestainedwith20µlmAbofvariousdilutionstoobtainatitrationcurveandtoidentifythesaturationpointanddetectionthreshold.Thefinalconcentrationoftheproductisthenadjustedtobeatleast3-foldabovethedetectionthreshold.Inadditionandtocontrollot-to-lotvariation,thegivenlotiscomparedandadjustedtofluorescencestandardswithdefinedintensity.

Storage:

Nordic-Mubiomonoclonalantibodyreagentscontainoptimalconcentrationsofaffinity-purifiedantibody.Forstabilityreasonsthismonoclonalantibodysolutioncontainssodiumazide.Thesereagentsshouldbestoredat2-8°C(DONOTFREEZE!)andprotectedfromprolongedexposuretolight.Stabilityofthereagent:Pleaserefertotheexpirydateprintedontothevial.Theuseofthereagentaftertheexpirationdateisnotrecommended.

References:

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Caution:

Forprofessionalusersonly.Thisreagentcontainssodiumazide.Toavoidthedevelopmentofhazardousconditions,reagentscontainingazideshouldbedilutedinrunningwaterpriortobediscarded.SimilartotheworkwithotherBIOLOGicalproducts,properhandlingproceduresarerecommended. Nordic的纯化的牛IgG可以在各种免疫测定（包括免疫扩散，免疫电泳，ELISA，Western印迹，斑点免疫结合测定（DIBA），血细胞凝集和细胞结合测定）中用作参考抗原，校准物，包被蛋白和封闭剂。该产品仅用于研究目的，并且用于体外测试，不适用于涉及人或动物的诊断或治疗程序。它可能包含有害成分。请参阅安全数据表（SDS）了解更多信息和正确的处理程序。根据当地法规处理产品残留物。此数据表的准确性尽可能合理，但Nordic-MUbio对本信息中的任何准确性或遗漏概不负责。