An Immunoassay for Dibutyl Phthalate Based on Direct Hapten Linkage to the Polystyrene Surface of Microtiter Plates 英文参考文献.docVIP

AnImmunoassayforDibutylPhthalateBasedonDirect HaptenLinkagetothePolystyreneSurfaceofMicrotiter Plates ChenxiWei1,ShumaoDing1,HuihuiYou1,YaranZhang1,YaoWang2,XuYang1*,JunlinYuan1* 1HubeiKeyLaboratoryofGeneticRegulationandIntegrativeBiology,CollegeofLifeSciences,HuazhongNormalUniversity,Wuhan,China,2CollegeofLifeSciences, FudanUniversity,Shanghai,China Abstract Background:Dibutylphthalate(DBP)ispredominantlyusedasaplasticizerinplasticstomakethemflexible.Extensiveuse of phthalates in both industrial processes and other consumer products has resulted in the ubiquitous presence of phthalates in the environment. In order to better determine the level of pollution in the environment and evaluate the potentialadverseeffectsofexposuretoDBP,immunoassayforDBPwasdeveloped. Methodology/PrincipalFindings:AmonoclonalantibodyspecifictoDBPwasproducedfromastablehybridomacellline generated by lymphocyte hybridoma technique. An indirect competitive enzyme-linked immunosorbent assay (icELISA) employingdirectcoatingofhaptenonpolystyrenemicrotiterplateswasestablishedforthedetectionofDBP.Polystyrene surface was first oxidized by permanganate in dilute sulfuric acid to generate carboxyl groups. Then dibutyl 4- aminophthalate,whichisananalogueofDBP,wascovalentlylinkedtothecarboxylgroupsofpolystyrenesurfacewith1- ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Compared with conjugate coated format (IC50=106ng/mL),thedirecthaptencoatedformat(IC50=14.6ng/mL)improvedassaysensitivityaftercarefuloptimization of assay conditions. The average recovery of DBP from spiked water sample was 104.4% and the average coefficient of variationwas9.95%.GoodagreementoftheresultsobtainedbythehaptencoatedicELISAandgaschromatography-mass spectrometryfurtherconfirmedthereliabilityandaccuracyoftheicELISAforthedetectionofDBPincertainplasticand cosmeticsamples. Conclusions/Significance: The stable and efficient hybridoma cell line obtained is an unlimited source of sensitive and specificantibo