Expression and purification of the recombinant outer membrane protein Tp0453 of Treponema pallidum and its characterization of immuno-competence-中国Betway体育网页登陆

摘要 TocloneandexpresstherecombinantoutermembraneproteinTp0453ofTreponemapallidumandtoanalyzetheimmuno-reactivityandimmunogenicityoftheexpressedprotein,theimmuno-dominantepitopeoftheTp0453wasamplifiedfromthecompletegenomeofT.pallidumbyPCR,subclonedintoexpressionvectorpQE32togeneratetherecombinantplasmidpQE32/Tp0453,thenexpressedinE.coliM15andanalyzedbySDS/PAGEandWesternblotting.ThefusionproteinexpressedwaspurifiedwithNi-NTAaffinitychromatography.Itsimmuno-reactivitywasassayedbyindirectELISA,andtheimmunogenicitywasdeterminedbyimmunizationwiththisfusionproteininNewZealandrabbits.Inthepresentstudy,afusionproteinofmolecularweightabout32kDawasobtained.AsdemonstratedbyWesternblotting,therecombinantproteincouldreactspecificallywithpositiveIgGseraofpatientswithsyphilis,andtheantibodiesagainstT.palliduminhumanseraweresuccessfullydetectedbyindirectELISA.BoththesensitivityandspecificityofELISAbasedontheTp0453fusionproteinaswere100%(30/30)whendetectedwithcontrolsera.IncomparisonwiththeresultsofIgGELISAwiththoseofTPPA.ItwasfoundthatthesensitivityofELISAwas96.8%andthespecificitywas100%.ThedifferenceofELISAandTPPAwasnotsignificant,andtheconcordanceofresultsbetweenELISAandTPPAwas98.2%.Inaddition,specifichumoralresponsescouldbeelicitedbyimmunizationwiththerecombinantfusionproteininNewZealandrabbitswithaspecificantibodytiterof1:1280after3successivedosesofimmunization.Theseresultsdemonstratethattheexpressedrecombinantfusionproteinshowsexcellentimmuno-competenceandprovidefoundationtodevelopaquickdiagnostickidappliedtodetectthepresenceofT.palliduminfections.

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Expression and purification of the recombinant outer membrane protein Tp0453 of Treponema pallidum and its characterization of immuno-competence

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