ELISA标准曲线制作方法

1、ELISA标准曲线制作方法一般而言，我们拟合ELISA标准曲线选用比较经典的CurveExpert1.3或者CurveExpert1.4软件。现在我们以CurveExpert1.4为例，对ELISA标准曲线绘制方法进行详述。Generallyspeaking,thestandardfittingcurveofELISAisbasedontheclassicsoftwareofCurveExpert1.3orCurveExpert1.4.NowwewillusetheCurveExpert1.4toillustratehowtoconstructtheELISAstandardcurve.1.点

2、击CurveExpert1.4，打开应用程序，截图界面如下：ClickCurveExpert1.4,andthenopentheapplicationprograms.Youwillseethefollowingscreenshot.fCurveExpert1.4FileEditDataInterpolateApplyFitToolsWindowHe-lpx|丫12x|丫1234567a91011121314151G17Fl-ankRegressionataPotCurveExpert1.4-UNREGISTEREDCOPYNUMUNTITLED.DAT2在X轴输入标准品的OD值，Y轴输入相应

3、的标准品的浓度，截图界面如下：InputthevalueofODontheX-axisagainsttheconcentrationofsamplesontheY-axis.Thefollowingscreenshotwillappearlikethis.3.单击上图界面中的Clicktheicon“showedontheabovescreenshotandyouwillseethefollowingpicture.CurveFinderCurveFird&reHamirie?yaurd曰t曰vndchoosesthebestpossiblecur?efit.Selectthemodelfam

4、ili&stoincludeinthecalculation.ModelFamilies审Poliinonnidls两LinearRegressions3.单击上图界面中的Clicktheicon“showedontheabovescreenshotandyouwillseethefollowingpicture.CurveFinderCurveFird&reHamirie?yaurd曰t曰vndchoosesthebestpossiblecur?efit.Selectthemodelfamili&stoincludeinthecalculation.ModelFamilies审Poliino

5、nnidls两LinearRegressions审ExponentialFarmiljJ1PowerLawFamily|7Yield-DensityModels冋Ori|妣可|7SigmoidalModelsPGrowthModels帀MiscellsrieousModels回UserModelsMaKimumDegreeofPolynomialto匚ansider:LilOKCarcelHelp4.单击上图界面中的ALLOFF按钮，出现如下界面ClicktheAllOffbutton,youcanseethefollowingscreenshot.CurveFiCurveFindr3bCur

6、yeFinderewaminesysurdataandchcase?thebestpossibleturvefit.Selectthemodelfamiliestoincludeinthecalculation.ModelFamilies厂Pcilynorriials厂LinearRegressionsI-ExponentialFamilyI-PswerLawFamily厂Yield-DensityModels|ZICln|AIIM厂SigmoidalModels厂GrowthModelsI-MiscellaneousModelsI-UserModelsMaMimumDegreeofPolji

7、namialtoConsider:OKCancelHelpIOKCancelHelpI5.我们选择SigmoidalModels这种曲线拟合模型进行ELISA标准曲线的拟合，点击OK,出现下图界面：WeselecttheSigmoidalModelstofittheELISAstandardcurve.ClickOK,andyouwillseethefollowingscreenshot.匾jLogisticModelI匾jLogisticModelPleasepresstherightmousebuttonforthegraphingfeaturesmenu.PressF1Forhelp_S

8、=U2.84G48G70r=0-99939327XAxis(units)注意：上图界面中的r值越高，即当r值越接近1时，所拟合的曲线就越好，所得到的数值可靠性也就也高。Pleasepresstherightmousebuttonforthegraphingfeaturesmenu.PressF1Forhelp_S=U2.84G48G70r=0-99939327XAxis(units)Pleasenotethatthehigherther-valueintheabovepicture(orthecloserther-valueisto1),themorereliablevaluesyouwill

9、get.在上图空白界面，按Ctrl键+L键，出现如下界面PleasepresstheCtrlplusLintheblankpartoftheabovepicture.Theinterfacewillappearlikethis.S=142.S4G48G7Oi=0.99939327Pleasepresstherightmousebuttonforthegr-aphingfeatur&smenu.PressF1forhelp.S=142.S4G48G7Oi=0.99939327Pleasepresstherightmousebuttonforthegr-aphingfeatur&smenu.Pre

10、ssF1forhelp.AtX=在上图中输入相应的OD值(X值)后，点击Calculate即可计算出待测物的浓度(Y值),待测物稀释了多少倍就在所测值的基础上乘以稀释倍数即最终浓度。AfterinputtingthecorrespondingOD-value(x-value),clickthebutton“Calculate.Theconcentration(Y-value)ofsamplescanbemeasured.Thefinalconcentrationcanbeobtainedbymultiplyingthethemeasuredfigurewiththetimesthesample

11、shavebeendiluted./Info点击所绘制的曲线图中左上方按钮，可以得到如下ELISA曲线拟合方程IInfoClickthebuttononthetopleftcornerinthepicture.ThenyoucangettheClickthebuttonXAxis(units)ELISAcurvefittingequationasfollows.匾；LogisticModelXAxis(units)点击上图中Copy即可得到上图界面中的曲线方程数据LogisticModel:y=a/(l+b*exp(-cx)CoefficientData:a=-1.19763443774E+004b=-8.45403498052E+001c=1.30213993104E+000Clickthebutton“cop