FOXP3isaforkheadtranscriptionfactorfamilymemberinvolvedinT-cellregulation,activationanddifferentiation.FOXP3hasbeenshowntobeamastercontrolgeneforthedevelopmentandfunctionofCD4+/CD25+regulatoryT-cells.InIHC,FOXP3hasbeenshowntobeaspecificMarkerforadultT-cellleukemia/lymphoma(1).Inmelanomaandinbreastandlungcancers,highnumbersofcirculatingregulatoryTcellshavebeenassociatedwithdiseaseprogression(2-5).Conversely,theinfiltrationofFOXP3+regulatoryTcellsintoinvasivetumorshasalsobeenreportedtobeassociatedwithsurvivalinavarietyofcancers(2-7).Incoloncancers,ahighfrequencyofFOXP3+infiltrateshasshowntobeapositiveindicator(6-7).PatientswithhighFOXP3expressioninCrohn’sdiseasehaveshownabetterresponsetoinfliximabtherapy(8).Inallographyrecipients,FOXP3celllevelsmayalsobeusefulinimprovingpost-transplantmanagement(9).
| ByLetter | F |
|---|---|
| IntendedUse | IVD |
| Format | Concentrate,Predilute |
| Clone | 86D |
| Volume | 0.1ml,1.0ml,6.0ml |
| Isotype | IgG1 |
| Localization | Nucelar |
| PositiveControl | Coloncancerandtonsil |
References:
1.RoncadorG,etal.FOXP3,aselectivemarkerforasubsetofadultT-cellleukaemia/lymphoma.Leukemia.2005Dec;19(12):2247-53.
2.GerberAL,etal.HighexpressionofFOXP3inprimarymelanomaisassociatedwithtumorprogression.BrJDermatol.2014Jan;170(1):103-9.
3.AliHR,etal.AssociationbetweenCD8+T-cellinfiltrationandbreastcancersurvivalin12,439patients.AnnOncol.2014Aug;25(8):1536-43.
4.LiuH,etal.Tumor-infiltratinglymphocytespredictresponsetochemotherapyinpatientswithadvancenon-smallcelllungcancer.CancerImmunolImmunother.2012Oct;61(10):1849-56.
5.TaoH,etal.Densityoftumor-infiltratingFOXP3+TcellsasaresponsemarkerforinductionchemorADIotherapyandapotentialprognosticfactorinpatientstreatedwithtrimodalitytherapyforlocallyadvancednon-smallcelllungcancer.AnnThoracCardiovascSurg.2014;20(6):980-6.
6.LingA,etal.TheintratumouralsubsiteandrelationofCD8(+)andFOXP3(+)Tlymphocytesincolorectalcancerprovideimportantprognosticclues.BrJCancer.2014May13;110(10):2551-9.
7.FreyDM,etal.Highfrequencyoftumor-infiltratingFOXP3(+)regulatoryTcellspredictsimprovedsurvivalinmismatchrepairproficientcolorectalcancerpatients.IntJCancer.2010Jun1;126(11):2635-43.
8.SloanS,etal.FOXP3+regulatoryT-cellcountscorrelatewithhistologicalresponseinCrohn’scolitistreatedwithinfliximab.PatholInt.2014Dec;64(12):624-7.
9.StenardF,etal.DecreasesincirculatingCD4+CD25hiFOXP3+cellsandincreasesinintragraftFOXP3+cellsaccompanyallograftrejectioninpediatricliverallograftrecipients.PediatrTransplant.2009Feb;13(1):70-80.
10.CenterforDiseaseControlManual.Guide:SafetyManagement,NO.CDC-22,Atlanta,GA.April30,1976“DecontaminationofLaboratorySinkDrainstoRemoveAzideSalts.”
11.ClinicalandLaboratoryStandardsInstitute(CLSI).ProtectionofLaboratoryWorkersfromOccupationallyAcquiredInfections;ApprovedGuideline-FourthEditionCLSIdocumentM29-A4Wayne,PA2014.
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看引物能不能特异性扩增，你可以检查一下扩增后的熔解曲线，一般单一的峰是特异性引物，如果有双峰那就是扩出来了杂带
上面较高的是样本曲线，下面较低的是未加样本（茎环引物和引物、探针都加入)
一般来讲，进行real-time qPCR MasterMix都是2×的浓缩液，只需要加入模板和引物就可以。由于real-time qPCR灵敏度高，所以每个样品至少要做3个平行孔，以防在后面的数据分析中，由于Ct相差较多或者SD太大，无法进行统计分析。通常来讲，反应体系的引 物终浓度为100-400mM；模板如果是总RNA一般是10ng-500，如果cDNA，通常情况下是1ul或者1ul的10倍稀释液，要根据目的基因 的表达丰度进行调整。当然这些都是经验值，在操作过程中，还需要根据所用MasterMix，模板和引物的不同进行优化，达到一个最佳反应体系。在反应体 系配置过程中，有下面几点需要注意：
1. MasterMix不要反复冻融，如果经常使用，最好溶解后放在4度。
2. 更多的配制Mix进行，减少加样误差。最好能在冰上操作。
3. 每管或每孔都要换新枪头！不要连续用同一个枪头加样！
4. 所有成分加完后，离心去除气泡。
5. 每个样品至少3个平行孔。
