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1、WesternBlot图片灰度分析–PhotoshopComparingtheintensityofbandsonaWesternblotcanbedoneinanumberofwaysusingsoftwarethatiscommonlyfoundonlabcomputersorfreelyavailablefordownload.Thefollowingdocumentoutlinessomeofthosemethods.Foralongtime,thede-factostandardforanalysisinlabsthatd
2、idn'twanttospringformulti-thousand$$commercialdensitometrysoftwarewasAdobePhotoshoporoneofthecompetingphotoeditingprograms.Allyoureallyneedinaprogramisafreehandselectiontoolandawaytomeasurethemeangrayvalueinsidetheselection.We'llstartwithAdobePhotoshopasanexample(youca
3、nfindmanyreferencesintheliteraturethatincludephrasessuchas"densitometrywascarriedoutinPhotoshop"inthemethodssection).Tostartwith,you'llneedtoscaninyourxrayfilmonaflat-bedscanner.Thiscanbeacheapconsumerunit,oramoreexpensivetransparencyscannerifyouhaveaccesstosuchabeast.
4、Youcanscanthefilmasagrayscaleimage,andsettheresolutiontoamediumvalue(300-400dpi).1.OpenthescannedimageinPhotoshop.2.UnderImage>Mode,checkthegrayscaleoptionifit'snotalreadyselected.Wedon'tcareaboutcolorinformation,onlygrayscaleinformation,sowecandiscardthecolorinformati
5、on.1.UnderImage>Adjustments,selectInvert(orpressCtrl+I).Nowthedarkpartsofthefilmarelight,andthelightpartsaredark.Thisisusefullater,asthehigh-expressionbands,whicharedarkonthefilm,willhavehighnumericalvalueswhenwemeasurethem.Whenphotoprogramsreportdarkness/lightnessvalu
6、es,thedarkpointshavevaluesnearzero,andthelightpointshavevaluesthatmaxoutat255.Theinvertedscan2.Choosethelassotoolfromthetoolpalette.3.Onthefirstband,usethelassotooltodrawalineallthewayaroundtheedgesofthefirstband.Thisiswhereyourjudgementcomesintoplay,determiningwhereth
7、eedgesofthebandare,andwhatissimplybackground.6.ForPhotoshopCS2/3:Ifthehistogramwindowisnotopenbydefault,gotoWindow>Histogramtoopenthehistogram.InnewerversionsofPhotoshop(CS2,CS3)thereisasmallarrowinacircleintheupperrightcornerofthehistogramwindow.Clickonthisandchoose'e
8、xpandedview'toshowthevaluesforyourselection.ForPhotoshopv.5+6:GotoImage>Histogramtodisplaythehistogramforthecurrentse
