A Molecular Assay for Sensitive Detection of Pathogen-Specific T-Cells 英文参考文献.docVIP

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A Molecular Assay for Sensitive Detection of Pathogen-Specific T-Cells 英文参考文献.doc

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A Molecular Assay for Sensitive Detection of Pathogen-Specific T-Cells 英文参考文献

AMolecularAssayforSensitiveDetectionofPathogen- SpecificT-Cells VictoriaO.Kasprowicz1,2,3*,JessicaE.Mitchell1,2,3,ShivanChetty2,3,PamlaGovender2,3 ,Kuan-Hsiang GaryHuang5,HelenA.Fletcher4,DanielP.Webster5,SebastianBrown5,AnneKasmar6 ,Kerry Millington7,CherylL.Day3,NompumeleloMkhwanazi3,CherylMcClurg1,2,3,FundisiweChonco3 ,Ajit Lalvani7,BruceD.Walker1,3,ThumbiNdung’u1,3,PaulKlenerman5 1RagonInstituteofMGH,MITandHarvard,HarvardMedicalSchool,Boston,Massachusetts,UnitedStatesofAmerica,2Kwazulu-NatalResearchInstituteforTBandHIV, Nelson R. Mandela School of Medicine, Durban, South Africa, 3HIV Pathogenesis Programme, Doris Duke Medical Research Institute, Nelson R. Mandela School of Medicine, Durban, South Africa, 4Nuffield Department of Medicine, The Jenner Institute, University of Oxford, Oxford, United Kingdom, 5Nuffield Department of Medicine, Oxford Biomedical Research Centre and James Martin School for 21st Century, University of Oxford, Oxford, United Kingdom, 6Division of Rheumatology, Immunology and Allergy, Brigham and Women’s Hospital, Harvard Medical School, Boston, Massachusetts, United States of America, 7Tuberculosis Research Unit, DepartmentofRespiratoryMedicine,NationalHeartandLungInstitute,ImperialCollegeLondon,London,UnitedKingdom Abstract Herewedescribethedevelopmentandvalidationofahighlysensitiveassayofantigen-specificIFN-cproductionusingreal timequantitativePCR(qPCR)fortworeporters-monokine-inducedbyIFN-c(MIG)andtheIFN-cinducibleprotein-10(IP10). WedevelopedandvalidatedtheassayandappliedittothedetectionofCMV,HIVandMycobacteriumtuberculosis(MTB) specificresponses,inacohortofHIVco-infectedpatients.Wecomparedthesensitivityofthisassaytothatoftheexvivo RD1(ESAT-6andCFP-10)-specificIFN-cElispotassay.Weobservedaclearquantitativecorrelationbetweenthetwoassays (P,0.001).OurassayprovedtobeasensitiveassayforthedetectionofMTB-specificTcells,couldbeperformedonwhole blood samples of fingerprick (50uL) volumes, and was not affected by HIV-mediated immunosuppressi