ELISA標(biāo)準(zhǔn)曲線制作方法

1、ELISA標(biāo)準(zhǔn)曲線制作方法一般而言，我們擬合ELISA標(biāo)準(zhǔn)曲線選用比較經(jīng)典的CurveExpert1.3或者CurveExpert1.4軟件?，F(xiàn)在我們以CurveExpert1.4為例，對(duì)ELISA標(biāo)準(zhǔn)曲線繪制方法進(jìn)行詳述。Generallyspeaking,thestandardfittingcurveofELISAisbasedontheclassicsoftwareofCurveExpert1.3orCurveExpert1.4.NowwewillusetheCurveExpert1.4toillustratehowtoconstructtheELISAstandardcurve.1.點(diǎn)

2、擊CurveExpert1.4，打開應(yīng)用程序，截圖界面如下：ClickCurveExpert1.4,andthenopentheapplicationprograms.Youwillseethefollowingscreenshot.fCurveExpert1.4FileEditDataInterpolateApplyFitToolsWindowHe-lpx|丫12x|丫1234567a91011121314151G17Fl-ankRegressionataPotCurveExpert1.4-UNREGISTEREDCOPYNUMUNTITLED.DAT2在X軸輸入標(biāo)準(zhǔn)品的OD值，Y軸輸入相應(yīng)

3、的標(biāo)準(zhǔn)品的濃度，截圖界面如下：InputthevalueofODontheX-axisagainsttheconcentrationofsamplesontheY-axis.Thefollowingscreenshotwillappearlikethis.3.單擊上圖界面中的Clicktheicon“showedontheabovescreenshotandyouwillseethefollowingpicture.CurveFinderCurveFird&reHamirie?yaurd曰t曰vndchoosesthebestpossiblecur?efit.Selectthemodelfam

4、ili&stoincludeinthecalculation.ModelFamilies審Poliinonnidls兩LinearRegressions3.單擊上圖界面中的Clicktheicon“showedontheabovescreenshotandyouwillseethefollowingpicture.CurveFinderCurveFird&reHamirie?yaurd曰t曰vndchoosesthebestpossiblecur?efit.Selectthemodelfamili&stoincludeinthecalculation.ModelFamilies審Poliino

5、nnidls兩LinearRegressions審ExponentialFarmiljJ1PowerLawFamily|7Yield-DensityModels冋Ori|妣可|7SigmoidalModelsPGrowthModels帀MiscellsrieousModels回UserModelsMaKimumDegreeofPolynomialto匚ansider:LilOKCarcelHelp4.單擊上圖界面中的ALLOFF按鈕，出現(xiàn)如下界面ClicktheAllOffbutton,youcanseethefollowingscreenshot.CurveFiCurveFindr3bCur

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

7、namialtoConsider:OKCancelHelpIOKCancelHelpI5.我們選擇SigmoidalModels這種曲線擬合模型進(jìn)行ELISA標(biāo)準(zhǔn)曲線的擬合，點(diǎn)擊OK,出現(xiàn)下圖界面：WeselecttheSigmoidalModelstofittheELISAstandardcurve.ClickOK,andyouwillseethefollowingscreenshot.匾jLogisticModelI匾jLogisticModelPleasepresstherightmousebuttonforthegraphingfeaturesmenu.PressF1Forhelp_S

8、=U2.84G48G70r=0-99939327XAxis(units)注意：上圖界面中的r值越高，即當(dāng)r值越接近1時(shí)，所擬合的曲線就越好，所得到的數(shù)值可靠性也就也高。Pleasepresstherightmousebuttonforthegraphingfeaturesmenu.PressF1Forhelp_S=U2.84G48G70r=0-99939327XAxis(units)Pleasenotethatthehigherther-valueintheabovepicture(orthecloserther-valueisto1),themorereliablevaluesyouwill

9、get.在上圖空白界面，按Ctrl鍵+L鍵，出現(xiàn)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=在上圖中輸入相應(yīng)的OD值(X值)后，點(diǎn)擊Calculate即可計(jì)算出待測(cè)物的濃度(Y值),待測(cè)物稀釋了多少倍就在所測(cè)值的基礎(chǔ)上乘以稀釋倍數(shù)即最終濃度。AfterinputtingthecorrespondingOD-value(x-value),clickthebutton“Calculate.Theconcentration(Y-value)ofsamplescanbemeasured.Thefinalconcentrationcanbeobtainedbymultiplyingthethemeasuredfigurewiththetimesthesample

11、shavebeendiluted./Info點(diǎn)擊所繪制的曲線圖中左上方按鈕，可以得到如下ELISA曲線擬合方程IInfoClickthebuttononthetopleftcornerinthepicture.ThenyoucangettheClickthebuttonXAxis(units)ELISAcurvefittingequationasfollows.匾；LogisticModelXAxis(units)點(diǎn)擊上圖中Copy即可得到上圖界面中的曲線方程數(shù)據(jù)LogisticModel:y=a/(l+b*exp(-cx)CoefficientData:a=-1.19763443774E+004b=-8.45403498052E+001c=1.30213993104E+000Clickthebutton“cop