Quantitative Analysis of Serum Procollagen Type I C-Terminal Propeptide by Immunoassay on Microchip 英文参考文献.docVIP

QuantitativeAnalysisofSerumProcollagenTypeI C-TerminalPropeptidebyImmunoassayonMicrochip ShoukiYatsushiro1,RieAkamine1,ShoheiYamamura1,MamiHino1,KazuakiKajimoto1,KaoriAbe1, HirokoAbe1,Jun-ichiKido2,MasatoTanaka1,YasuoShinohara3,4,YoshinobuBaba1,5,ToshihikoOoie1, MasatoshiKataoka1* 1HealthResearchInstitute,NationalInstituteofAdvancedIndustrialScienceandTechnology(AIST),Japan,2DivisionofMedico-DentalDynamicsandReconstruction, DepartmentofPeriodontologyandEndodontology,OralandMaxillofacialDentistry,InstituteofHealthBiosciences,UniversityofTokushima,Tokushima,Japan,3Faculty ofPharmaceuticalSciences,UniversityofTokushima,Tokushima,Japan,4InstituteforGenomeResearch,UniversityofTokushima,Tokushima,Japan,5Departmentof AppliedChemistry,GraduateSchoolofEngineeringNagoyaUniversity,Nagoya,Japan Abstract Background: Sandwich enzyme-linked immunosorbent assay (ELISA) is one of the most frequently employed assays for clinical diagnosis, since this enables the investigator to identify specific protein biomarkers. However, the conventional assayusinga96-wellmicrotitrationplateistime-andsample-consuming,andthereforeisnotsuitableforrapiddiagnosis. Toovercomethesedrawbacks,weperformedasandwichELISAonamicrochip. MethodsandFindings:Themicrochipwasmadeofcyclicolefincopolymerwithstraightmicrochannelsthatwere300 mm wideand100mmdeep.FortheconstructionofasandwichELISAforprocollagentypeIC-peptide(PICP),abiomarkerfor boneformation,weusedapiezoelectricinkjetprintingsystemforthedepositionandfixationofthe1stanti-PICPantibody onthesurfaceofthemicrochannel.Aftertheinfusionofthemixtureof2.0 mlofperoxidase-labeled2ndanti-PICPantibody and 0.4ml of sample to the microchannel and a 30-min incubation, the substrate for peroxidase was infused into the microchannel; and the luminescence intensity of each spot of 1st antibody was measured by CCD camera. A linear relationship was observed between PICP concentration and luminescence intensity over the range of 0 to 600ng/ml (r2=0.991),andthedete