final test of 三维显微术

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1、FinaltestTofindevidenceofthefourquestions,Idesignedanexperimenttoverifythefourprocessesmainlyintwosteps.Oneisusingscanningelectronmicroscopy,theotheriscryoET.SamplepreparationChemicalfixation:topreserveandstabilizetheirstructure.Fixationisusuallyperformedbyincubationinasolutionofabufferedchemicalf

2、ixative,suchasglutaraldehyde.Appropriatesamplesize:samplemustbeofappropriatesizetofitthespecimenchamberandgenerallymountedrigidlyonaspecimenholdercalledaspecimenstub.Coating:useconventionalSEMsputtercoatingthemajorprocessesasthefigureshows.First:Theconventionalsputterchamberispumpedwitharotarypump

3、untilthevacuumlevelisbeyondhalfscale.Second:Leakargongasintothechambertofloodthechamber,allowthevacuumtorecover.Third:Applythelowestvoltagethatwillallowaplasmatostrike.Note:incaseofsmallparticlesitwillbegoodtotiltitbyabout45℃.SEMscanningThefirstthreequestionscanbeconcludedfromthepicturebelow.Liket

4、hepictureshows,1.thesurfaceofthecellsandthenanoparticlemaylookgreywithmanybumpsandpits.Buttheimagecanbegreenifusingthelightmicroscopyfornanoparticle.2.Thenanoparticlecanenterthecellontheconditionthatthereceptorshaveidentifiedthespecificsolublesubstance.Thenanoparticleenteringthecellbyfusionorbyend

5、ocytosisorothermethodisdeterminedbytheimagestakenfromSEM,eachofthemhasthepossibility.StructureofthenanoparticleBrieflysamplepreparation:nano-particlesarespreadintothinfilmsacrossEMgridsplunge-frozeninliquidethanetransferredintoanelectroncryomicroscope.Note:Plunge-freezingpreventsicecrystallization

6、,immobilizingtheproteinsandothercellularstructuresintheirnativestatesandlocations.ThetheorylikethepicturesshowbelowProjectionreconstructionback-projectionOperation:imagediterativelywhilebeingtiltedincrementally.Thenrecordingaseriesofimagesatdifferentanglesallowsafull,three-dimensional(3D)reconstru

7、ctionofthespecimentobecalculated.Makingmodel:afterthe3Dreconstruction,Icanbuildamodeltoreflectthestructureofthenano-particle.Fromthemodel,apparentlycangetsomeinformationaboutthesizeandshapeofthenano-particle.Butw