fall 2016 newsle-er - notre dame integrated imaging facility · fall 2016 newsle-er ... in vivo...
TRANSCRIPT
NDIIFNewsBriefs p. 2
AdvancedElectronMicroscopyCore p. 4
InVivoImagingCore p. 11
OpCcalMicroscopyCore p. 17
HistologyCore p. 27
NDIIFPolicies p. 28
Fall2016Newsle-er
This newsleJer summarizes current capabiliCes and includes graphsshowing usage trends to date for 2016. The NDIIF operates as a rechargefacilityandrevisesuserfeeseveryJuly1.
QuesCons about NDIIF policies should be directed to the Director or anymemberofthesteeringcommiJee.FortechnicalandschedulingquesCons,seetherelevantNDIIFstaffmemberslistedonthefollowingpages.Contactnumbers and addiConal informaCon can be gained by visiCnghJp://www.imaging.nd.edu.
SincerelyProfessorBradleySmith,[email protected]
TableofContents
1
NDIIFNewsBrief
1. MidwestImagingandMicroanalysisWorkshopatNotreDame:The
thirdannualworkshop,heldMay2016,wasverysuccessfulwithafocusonelectronbeamtechnologies.TheeventwillberepeatedmidMay2017.
2. NDIIFAwardsforBestImagingPublicaIons2015:
• TheBestElectronMicroscopyImagingPublicaConAwardfor2015wasawardedtoSarahFathipour,agraduatestudentwithProfessorAlanSeabaughintheDepartmentofElectricalEngineering.
• TheBestBiologicalImagingPublicaConAwardfor2015wasawardedtoDr.ManuelaLahne,AResearchAssistantProfessorandacollaboratorwithProfessorDavidHydeintheDepartmentofBiologicalSciencesandtheCenterforZebrafishResearch.Thesubmissiondeadlineforthe2016awardsismidFeb2017.
3. CTSICoreFacility:AffiliaConwithCTSIasacoreresearchfacilitywasrenewed.FormoreinformaConpleasevisit:hJp://www.indianactsi.org
4. InVivoImagingCore:acquiredaUniversalLaserCuJerVLS6.60–
LasercuJer/engraver.
5. ElectronMicroscopyCore:acquiredaDENSheaCngandbiasingholderforatomicresoluConimagingofin-situheaCng.
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Director:Dr.BradleySmith,COSSteeringCommittee:Dr.KevinVaughan(Chair),COSDr.HollyGoodson,COSDr.CrislynD’Sousa-Schorey,COSDr.KenKuno,COSDr.GaryBernstein,COEDr.GlenNiebur,COEDr.PaulMcGinn,COEOf1iceManagerTheresaBollingerDirectorofBiologicalImagingDr.W.MatthewLeevy,BiologicalImagingAssistantDirectorSarahChapman,LabMangerCellMicroscopyWilliamArcher,DirectorofElectronMicroscopyDr.AlexMukasyanLabManagerSEM,FIBTatyanaOrlovaTEMProgramDirectorDr.SergeiRouvimovMicroprobeProgramManagerDr.IanSteele
NOTREDAMEINTEGRATEDIMAGINGFACILITYORGANIZATIONALPLANFY16
NDIIFMissionStatementEstablished in 2008, the NDIIF provides an integrated suite ofsophis<catedmicroscopes and imaging sta<ons that enable the expertuserstoa@ackthemostcomplexmodernresearchproblemsand,equallyimportant, the resident professional staff (technicians and researchspecialists)toguidethenon-expertusers.
3
ADVANCEDELECTRONMICROSCOPYCORE
AboutUs:TheAdvancedElectronMicroscopyCoreintegratesasuiteofsophisCcateddevices that enable theexpertusers to aJack themost complexmodernresearch problems frommicron to Angstrom spaCal resoluCon and theresidentprofessionalstafftoguidethenon-expertusers.Thecorecontainsauniquebundleofthreestate-of-the-artFEItoolsandincludes:
• Magellan 400, Digital Field Emission Scanning Electron Microscope(FESEM),recordhighspaCalresoluCon0.6nm;• Helios NanoLabTM DualBeam 600, SEM/Focus Ion Beam (FIB)WorkstaCon that is capable of nano-prototyping, nano-machining, nano-analysis,andadvancedTEMsamplepreparaCon;• EVO50Zeiss,environmental SEMwithPelCerStage,allowsoperaConunderhighandlowvacuumcondiCons.• Titan 80-300, Transmission Electron Microscope (TEM) enables sub-Angstrom,atomic scalediscoveryandexploraCon inbothTEMandSTEMmodesoverawiderangeofmaterialsandoperaCngcondiCons.• JEOL2011,TransmissionElectronMicroscope(TEM)80-200kV,0.14nmresoluCon,workhorseformaterialsandbiologicalsamples.Researcherscanbetrainedtouseourdevices,orservicecanbeprovidedbyexperiencedstaff.ForgeneralquesConsabouttraining,orcontractedimageacquisiConandanalysis,contact,Dr.AlexanderMukasyan574-631-9825oramoukasi@nd.eduForquesConsaboutTEMcontactDrSergeiRouvimov574-631-0226orsergei.rouvimov.1@nd.edu
Dr.AlexanderMukasyanResearchDirector
Mrs.TanyaOrlovaResearchTechnician
!
Dr.SergeiRouvimov,TEMProgramDirector
Dr.IanSteele,MicroprobeProg.Director
SEM/FIBWorkstaOon:HeliosNanoLabDualBeam600(FEI):
SpecificaOons:ElectronopOcs:-Resolu<on:0.9nm@15kV,1.4nm@1kV-Detec<on:in-lensSEandBSEIonopOcs: Sidewinder™fieldemissionfocusedionbeamopCcswithliquidGalliumionemiJer;-Resolu<on:5.0nm@30kV-Detec<on:CDEMdetector
TheHeliosNanoLab™WorkstaIoniscapableof:• nano-prototyping• nano-machining• nano-analysis• advancedTEMsamplepreparaCon
Cross-secConofhighlyporous(93%)metalbulkalloy(Prof.Mukasyangroup)
ApplicaOon:• PaWerning:Simultaneousimagingandpa@erningwithend-pointdetec<onthroughReal-TimeMonitor;• AutoFIB;• PlaOnumDeposiOon;• SelecOveCarbonMill;• EnhancedEtch;• TEMsamplepreparaOon:-AutoTEMG2;OmniprobeAutoProbe200.2DiagnosOcs:-EDS&EBSD-PegasusPackage,EDAX
CrosssecConofamesasarrayforlayerthicknessmeasurementandstructureprofilecontrol(Prof.TangfeiLuogroup)
EBSDfortexturedAlalloy(Prof.S.Songroup,PurdueUniversity)
FESEM:Magellan400(FEI)TheMagellan400isafullydigitalFESEMwithSchoJkeyfieldemiJersourcemountedontheNGhot-swapgunmodulethatprovidesrecordhighspaCalresoluCon:0.6nm@15kV,0.9nm@1kVFeaturesandspecificaOons:-in-lensSEandBSEdetecConspeciallydesignedforhigh-resoluConimagingatbothhighandlowkV’s;-Everhart-ThornleySEdetectorforSEdetecCon.;-AnintegratedIRCCDcameraforin-chamberviewing;- RetractableAnnularSTEMDetectorenablesscanningtransmissionimaginginbrightfield,darkfieldandhigh-angledarkfieldmodes.EnergyDispersiveX-RaySpectrometer(EDS)Bruker-SLEWWindow,detecConofBoronandup;-energyresoluIon123eV(MnKα,0-100,000cps);-Elementalmappingandmore
TheMagellanXHRSEMallowssampleimagingatextremelylowbeamenergies(<100eV),avoidingchargeeffectatnon-conducCvenano-scalesurfaces.
Aring-diskelectrodearrayfabricatedonglassbynanospherelithographyandmulC-stepRIEetching(ChaoxiongMa)
Asteelexposedto1.5MeVelectronbeamirradiaCon(SlavicaGrdanovska)
StaphylococcusaureusinteracCngwithepithelialcells(TrevorKane)
OlicellwithemergingM13bacteriophage.(MicheleCostanCno,IUSB)
EnvironmentalSEM:EVO50LEO(CarlZeiss)TheEVO50ZeissenvironmentalSEMequippedbyPelCerStage,whichallowstoworkbothunderhighandlowvacuumcondiCons,hasthefollowingspecificaCon:
Resolution 2.0nm@ 30kV (SE with LaB6 option )
Acceleration Voltage 0.2 to 30 kV
Magnification 35x to 1,000,000x
Field of View 6 mm at the Analytical Working Distance (AWD)
EDX-ray Analysis Oxford Instrument, resolution 133 eV
Available Detectors
• SE in HV - Everhart-Thornley • SE in VPSE • BSD in all modes - quadrant semiconductor diode
Chamber Approx. 365 mm (dia.) x 255 mm (h)
5-Axes Motorized Compucentric Specimen Stage
• X = 100 mm (+50mm, -50mm) • Y = 125 mm (+65 mm, - 60 mm) • Z = 55 mm (35 mm motorized) • T = 00 - 900 • R = 3600 (continuous) Stage control by mouse or optional joystick and control panel
Image Processing Resolution: Up to 2304x1024 pixel Signal acquisition by integrating and averaging
Image Display Single flicker-free XVGA monitor with SEM image displayed at 1024 x 768 pixel
System Control Smart-SEM with Windows, operated by mouse, keyboard and optional control panel.
Element Weight% Atomic% C K 3.10 14.48 O K 0.73 2.56 P K 13.35 24.19 Ga K 41.48 33.40 As L 19.76 14.81 In L 21.58 10.55
Quantitative results
Wei
ght%
0
10
20
30
40
50
C O P Ga As In
ApplicaIons:UserfriendlyandwitharelaCvelylowuserfee,theEVO50ZeissisanaJracCveopConforrapidscreeningofsamplesmicrostructures.ItisorenusedforEDXanalysiswithspaCalresoluConof1µm
EDXspectraanddataofelementalanalysisinthespotofGa-basedmonocrystal
FeaturesandspecificaOons(at300kV):EnergySpread-0.7eVPointResolu<on-0.2nmInforma<onlimit-<0.1nmSTEMResolu<on-0.136nm
TheTitan80-300microscopeincorporatesanovelplasormthatallowsulCmatestability,performanceandflexibility:• HighResoluCon(HR)TEMmode• HRScanningTEM(STEM)mode• HRElectronEnergyLossSpectroscopy(EELS)• HREnergyDispersiveX-Ray(EDX)
TEM:Titan80-300(FEI)
ApplicaOon• TEMimagesandElectronDiffracOon• HighResoluOonTEMimagesatAngstromresoluOon• HighthroughputSTEMmodeatAngstromresoluOon• EDX-composiOonalanalysisatnm-scale• ElectronEnergyLossSpectroscopy
EELSallowstoobtainelementalandchemical(suchasvalenceandthecoordina<onofspecificatoms)informaOonwithnanometerresoluOon.HAADFSTEMimagesofgradedAlGaNlayers(incross
secCon)showingbothplanarspontaneoussuper-laucesand3DcomposiConalfluctuaConsintheAlGaNlayer.InsertedisEDSscanlinesshowingvariaConofAlandGaingradedAlGaNlayer(lerimage).S.Rouvimov,S.M.Islametal.“TEMAnalysisofStructureandComposiConalFluctuaConsinMBEgrownAlGaNStructuresforDeep-UVPhotonics”,EMC2015
(a)SchemaCcand(b)falsecoloredTEMcross-secConoftheAg/HfO2TSdevice.InsetshowsAg/HfO2/p+SiinterfacethatenablesthresholdswitchingacCon.Thecross-secConistakenalongx-y.(c)EDXscanoftheinterface.NikhilShukla,BenGrisafe,etal(groupofSumanDa-a),“Ag/HfO2basedThresholdSwitchwith107SelecCvityand100μAON-currentforCross-PointSelectorandSteep-slopePhase-FETApplicaCon”,2016IEEEInternaConalElectronDevicesMeeCng,SanFrancisco,CA,2016.
V.Kanzyuba,S.Dong,etal(groupofProf.J.Furdyna),“StructuralproperCesofSnMnSe:anew2DmagneCcsemiconductorwithpotenCalforspintronicapplicaCons”,MM2016;S.Dong,etal“RoomtemperatureweakferromagneCsminSn1−xMnxSe22Dfilmsgrownbymolecularbeamepitaxy”,APLMATERIALS4,032601(2016)
TechnicalSpecificaIons•OperaCngvoltageof80–200kV•HRPolePiecewithresoluConof0.14nm.•MagnificaConof50x–1,500,000•JEOLsingleCltholder•Gatan636DoubleTiltholder•Gatan622TVcamera•CCDcamera*•OxfordINCA30mm2LN2detector
TEM:JEOL2011
Low-andhigh-magnificaConTEMimagesof(a,c)CdSand(b,d)CdS/NiNSs.InsetsgivethecorrespondingensembleSAEDimages(a,b)andahigh-resoluConTEMimageshowingbasalplanelaucefringes(toprightin(c))aswellasaNSsideview(boJomlerin(c)).EnsembleEDXSspectraof2.16nm(e)CdSand(f)CdS/NiNSs.).MaksymZhukovskyi,etal(groupofProf.Kuno)“EfficientPhotocatalyCcHydrogenGeneraConfromNiNanoparCcleDecoratedCdSNanosheets”,ACSCatal.2015,5,6615−6623
!
This “cerCfied” used instrument (2001) isfully operaConal. For quesCons about TEM,trainingandapplicaCons,please,contact DrSergeiRouvimov,233SCnson-RemickHallE-mail:[email protected]:574-631-0226(office)
Atomicmodels(top)andHRTEMimages(boJom)ofSnS2toCu2SnS3.InsertedarediffracConpaJerns.YuanxingWang,etal(groupofProf.K.Kuno),“TransformingLayeredtoNonlayeredTwo-DimensionalMaterials:CaConExchangeofSnS2toCu2SnS3”,ACSEnergyLeJ.2016,1,175−181
TEMImagesofLargeUnilamellarvesicleswithandwithoutprotein.Doestheproteinbendmembranes?KristenA.Johnson(groupofProf.R.V.Stahelin)
ElectronProbeAnalyzer:CamecaSX-50
TheCamecaSX-50canprovidehighaccuracychemicalanalysesofawidevarietyofpolishedsamples(e.g.minerals,rocks,meteorites,metals,glasses,cements,ceramics,etc.)withmicronresoluCon.ElementalimagescanbeobtainedusingeitherwavelengthofenergydispersivedetectorsinaddiContoBSEimages.
2 mm
Polished section
Location of sulfate deposition during discharge by imaging polished sections
of battery plates: Bright areas are PbSO4 formed during discharge.
Surface only – prevents acid reaction – battery fails
Surface and interior
Sulfate interior – good battery
Imaged areas from different batteries.
Chemicalanalysisandelementalimagingonamicronscale
Example:ApplicaIontoba-eryfailure
INVIVOIMAGINGCOREAboutUs:TheNotreDameInVivoImagingCoreprovidesanon-invasiveapproachtoobservevariousdiseaseandbiologicalcondiConsinlivingmice.Thisfacilitycurrentlyhasteninstrumentsundermanagement,witheightmodaliCesavailablefordirectuseraccess.First,theIVIS®LuminaenablesthesensiCvedetecConofbioluminescencefrommammalianandbacterialcellsthatexpressluminescentgeneCcreporterconstructs.Next,theBrukerXtremeisuClizedtoimagecellsthatexpressfluorescentgeneCcreporterslikeGFPorRFP,andalsodetectthebiodistribuConofnear-infrared(NIR)probesandnanomaterials.Further,theXtremeoffershighresoluCon,highspeedplanarX-rayforanatomicalandradiographicimaging.TheXtremealsooffersplanarscinCgraphicimagingofawiderangeofradiolabeledspecies.InaddiContoplanaropCcalandX-rayimagingimaging,thefacilityoffersnuclearimagingwithPositronEmissionTomography(PET),SinglePhotonEmissionComputedTomography,inaddiContoanatomicalimagingwithX-rayComputedTomography(CT).NuclearimagingmodaliCesareenabledbythestate-of-the-artAlibraTrimodalPET/SPECT/CT.AsuiteofX-rayComputedTomographyequipmentincludestheScanCoVivaCT80(smallFOV,highres),NeurologicaCereTom(largeFOV,lowres)andAlbira(medFOVandmedres).NuclearprobesarecommerciallyavailablethroughlocalvendorslikeSpectronMRC(PET)andCardinalHealth(SPECT)toenableaccesstocriCcalapplicaConareasinheart,tumor,lung,andbone.TomographicanatomicalimagingofsorCssuesisprovidedbyanICONMRI.Finally,theInVivoCorealsooffersrapidbodycomposiConanalysisofFatandLeanCssueweightsusinganEchoMRIsystem.OurgoalistotrainandempoweruserstoindependentlyuClizethefacilityanditsresources.ContacttheDirectororAssistantDirectorofBiologicalImagingforassistance
Prof.MaJhewLeevyDirector,BiologicalImaging
SarahChapmanAssistantDirectorofBiologicalImaging
INVIVOIMAGINGCORE
IVISLumina(Twounitsoncampus)InVivoBioluminescence
BrukerXtreme(Twounitsoncampus)InVivoFluorescencePlanarX-rayPanarScinCgraphy
ICONMRIDemothru2016AnatomicalimagingofsorCssue
KeyEquipment:
EchoMRIBodyComposiConAnalysis
INVIVOIMAGINGCORE
BrukerAlbiraPositronEmissionTomography(PET)SinglePhotonEmissionComputedTomography(SPECT)X-rayCT
KeyEquipment:
NeurologicaCereTomLargeBoreX-rayCT
ScanCoVivaCT80HighResoluConX-rayCT
MARSMedipixSpectralX-rayCT
Tumor Imaging - Time-course of healthy lung volumedegradaIonduring breast tumormetastasis. Calli Davison oftheSchaferLabisstudyingbreastcancermetastasisinlivingmiceusing non-invasive X-ray computed tomography on the AlbiraPET/SPECT/CT. Two cohorts of mice were injected with eitherbreast cancer cells, or saline (control), andwere imaged over aperiod of six weeks. The lung Cssue volume was segmented(shownaboveinpurple)andquanCfiedforeachgroup. ThelungdestrucConcausedbytumorgrowthisreadilyapparentfromtheinvivoimages. TheSchaferlabiscurrentlyusingthistechnologytostudythemetastaCcproperCesofbreastcancercelllineswithalteredexpressionlevelsoftheenzymecatalase.
ImagingBrainStroke–Prof.RashnaBalsaraisusingPositronEmissionTomography(PET)toimagebrainmetabolisminratswithcerebralischemia.TheimageshowstheCT(anatomicalmap,ler),PET(center)andPET-CToverlay(right)ofarat24hoursarerastrokeevent.
ALBIRAPET/SPECT/CT
TetramodalSPECT-PET-CT-MRIofaSingleMouse–TheLeevylabhasdevelopedanewmethodfortetramodaltomographicimagingofmice.HereweshowonemouseimagedbyCT(X-ray),MRI(SorTissue),PET(BoneScanwithNa18F),andSPECT(LungperfusionscanwithMAA-99mTc.
WK2
WK4
WK6
ImagingtheAnatomicalFeaturesofSpecimensRangingfromBeetlestoRats–X-rayCTmaybeusedtoimagetheinternalstructuralfeaturesof
SCANCOHIGHRESOLUTIONX-RAYCOMPUTEDTOMOGRAPHY(CT)
ratsorotherspecimenslikebeetles.ApplicaConsincludebonedensityimaging,anatomicalimaging,andphenotyping.
ALBIRAPET/SPECT/CTcont.EvanDoneyoftheLeevyLabisdevelopingnovelmethodsthatcombinepreclinicalimagingandaddiCvemanufacturing.RatsscannedwiththeAlbiraImageStaConX-rayCTmodality(top)wererenderedandeditedasstereolithographsandprintedasphysicalmodelsboJomusinganumberof3DprinCngplasorms,includingShapeways.com(boJom)
SorCssuesegmentaCon,prinCnginmulCplecolorsandmodelcustomizaConarealsopossiblewiththisnewlydevelopedmethod.TheIntegratedImagingFacilitycurrentlyhastwo3Dprintersandisimprovingandexpandingthesemethodstothemicroscopicleveltoincludehumandata,aswellascellsandorganelles.
FluorescenceImagingofBoneRemodeling–TheMSFXspecializesinthedetecConoffluorescentprobesinlivingmice.AtrightisamontageofX-ray,fluorescence,andoverlayofamouseinjectedwithOsteosense-750,aprobeforboneremodeling.ImagegeneratedbyundergraduateresearchersintheLeevyLab.
DynamicImagingofP.aeruginosaSwarmBehavior-ResearchersfromthelabofProf.JoshuaShroutareperformingCmelapsefluorescenceimagingGFP-expressingbacteriatomonitortheirgrowthandswarmkineCcs.AtlerisasingleframefromathreedayCmecourse,inwhicha“fire”colorscaleisusedtoindicatedtheintensityofGFPsignalcomingfromdifferentregionsontheplate.
BrukerMULTISPECTRALFXIMAGESTATION
OpIcalImagingofRFPOvarianCancerMets–TheCowden-DahllabisusingRFP-expressingovariancancercellstotrackandquanCfytumormetastasisduringexvivoimaging.TheimageatrightshowsaphotoofanIPregionofamouse(ler),theRFPfluorescenceimage(center),andanoverlayofboth.
InVivoImagingofaBioluminescentSalmonellaInfecIon–ResearchersfromtheSmithLabhavedevelopedinvivomodelsofinfecConusingbioluminescentbacteria.Thesebacteriaareengineeredtoemitlight,thusfacilitaCngtheirdetecConusingalightsensiCveCCDchip.BLIemissionreportsonthehealthoftheseinvadingcells,anddrugtherapymaybenon-invasivelymonitoredasadecreaseinlightemanaCngfrominfectedCssue.
IVISLUMINA
MulImodalProstateTumorImaging–AcollaboraConbetweenresearchersfromtheSuckowLab,LeevyLab,andDr.BrianRabinovichfromMDAndersen,hasproducedaprostatetumorcelllinewithdualbioluminescentandfluorescentgeneCcreporters.Fromlertorightisasub-Qtumorwithsignalcapturedinphotographic,mCherryfluorescence,exogenousMMPSense750probefluorescence,Cerenkovluminescence(FDG),andPET(FDG)modaliCes.
Photo mCherry MMPSense750 Cerenkov PET
OPTICALMICROSCOPYCOREAboutUs:TheOpCcalMicroscopyCore(OMC)givesresearchersaccesstohighendresearchmicroscopesallowingthemtoimage,eitherfixedorlive(inenvironmentallycontrolledchambers),fluorescentlylabeledcellsandCssuesthataremountedonslidesorinpetridished.Oursystemsletsusersworkwithuptofour(separatechannels)fluorescentmarkersintheemissionspectrarangingfromDAPIintheUVwavelengthstothefar-redfluorophoresnearingtheIRwavelengths.ThecontrollingsorwaresgivethemtheopportunitytoacquiresingleinformaCveimagesaswellasz-seriesimagesforthree-dimensionalreconstrucCons.ResearcherscanbetrainedtouseourdevicesinasliJleastwohours.TheCoreislocatedinSuite007inthebasementofGalvinLifeSciencesBuilding.ForconsultaCons,schedulingoftraining,orforcontractedimageacquisiCon,contacttheLabManager,BillArcher574.631.5443([email protected]).
BillArcherLabManager
OpCcalMicroscopyCore
ApplicaIons:geneCcallyencodedfluorescentproteins–anCbodies-fluorescentmolecularprobes–singlecellsandmulClayeredCssues–changesinCssuearchitecture–Cme-lapsemovies–movementandgrowthofsub-cellularorganellesandmacromolecules–fluorescentionicgradientreporters–imagingoffixedandlivingsystems:bacteria,proCsts,Drosophila,zebrafish,allculturedcelltypes–transparentnon-livingsamples
Equipment:NikonAZ100Marco/ZoomScope
TheNikonAZ100isamulC-purposemacrozoommicroscopethathasmulCplemethodsforonetoobservesamples(fixedslides,tosmallorganisms)eitherinbrighsield(toplitorbacklit),NomarskiDIC,orepi-fluorescence.ItcoversawiderangeofmagnificaCons,from10xto320xallowingtheusertoimagethesamesamplefrommacrotomicroobservaConsaswellascreatemoviesoflivesamples.
NomarskiDIC
Brighlield/BacklitBrighlield/Toplit
Above:ImagesbyJackieinDr.MichaelPfrender’slablookingatdevelopmentalstagesinthewaterfleaDaphnia
Below:ImagesbyChrisKegelmanworkinginDr.JoelBoerckel’slab
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Mouseembryoribcage Mouseembryo
skull
Mouseembryovertebra
NikonEclipse90iWidefieldFluorescentMicroscope
TheNikonEclipse90iisaneasy-to-usestandarduprightbrighsield/fluorescentresearchmicroscopeequippedwithtwocameras(onecolorandonemonochromaCc)andiscapableofcapturingsingleimagesormovies,ofwholeCssues,smallorganisms,etc.witharangeofobjecCvelensesfrom4xupto100x.
DrosophilaembryoDAPIwavelength Drosophilaembryo
FITCwavelength
Below:ImagesbyTsuyoshiTokusumiworkinginDr.RobertSchulz’slab
Above:ImagesbyBillArcher
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DiatomstakenwithmonochromaIccamera
BloodvesselsseeninasecIonofahumanuterus
GEHealthcareDeltaVisionDeconvoluIonMicroscopeTheDeltaVisionimagingsystemisaninvertedmicroscopethatallowstheusertocapturewidefieldepi-fluorescenceimages,andwiththedeconvoluConalgorithmredirectalloutoffocuslighttobecomeinfocusforsharpimages.Itcandothiswithfixedcellsand/orCssues,butitsgreateststrengthisitsabilitytoobtainandanalyzelong-termCme-lapsethree-dimensionalimageswithaminimumofphoto-damageonlivecellsorsmallorganismswiththeaidofitsenvironmentalchamber.TheimagescanbecapturedwithaCoolSnapHQ2cameraforhighresoluConimagesoranEMCCDCascadeII/512high-sensiCvitycameraforsampleswithlowlight.
ImagingofSubcellularStructures–Theimagesbelowshowcellsduringthemetaphaseperiodofcelldivisiontoexaminethestructure,assembly,andbehaviorofkinetochoresinliveculturedCOS-7cells,researchbeingdonethelaboratoryofDr.KevinVaughan.
Chromosomalarchitecture–offluorescentlymarkedchromosomesfromthegenomeofMalpighiantubulecellsofthemosquitoCulex.ThisworkwasdoneinDr.FrankCollinsLaboratory
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Andor/NikonSpinningDiskConfocalMicroscope
AndorTechnologyandNikonInstrumentsworkedtoproducethisSpinningDisk,livecellconfocalmicroscope.Thissystemisdesignedtoimagethinsampleswithlowemissionfluorescenceaswellascapturecellulareventswithminimalphoto-damagetothecells.ThissystemisalsocapableofpreformingthemodaliCesofFRET,FRAP,andTIRF.
Forareadablesummaryofspinningdiskmicroscopy,see:hJp://cshprotocols.cshlp.org/cgi/content/full/2010/11/pdb.top88
Recordingcytoskeletalbehavior–Dr.HollyGoodsonobtainsimagesofliveCOS-7cellsoverCme.UsingFluorescenceRecoveryArerPhoto-bleaching(FRAP)(redcircle)modalityshestudiestherealCmebuildingofthefluorescentlylabeledmicrotubulecytoskeletonaswellasthemovementofintracellularorganellesassociatedwiththecytoskeleton(greenarrowhead).
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ObservaConofmembraneandnearmembranephenomenawithTIRFmodality(CourtesyofD.ToomteoftheYaleUniversityofMedicineanAndorUSA)
NikonC-2LaserScanningConfocalMicroscope
The Nikon C-2 confocal microscope is a simpleoperaConal confocal setup on a Nikon Ni-E uprightresearch microscope and can be considered the liJlesister of the Nikon A1-R/MP confocal system. Thisallowsresearcherstoimageandanalyzetheirfixedcellsor Cssues from above, when an inverted scope is notpracCcalfortheirsamples.
A3-DmodelrenderedfromaCmelapse,z-seriesacquisiConlookingforcellmovementbetweenlayersofalivezebrafishreCnabyDr.ManuelaLahneworkinginDr.DavidHyde’sLaboratory.
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MolecularProbeslideofBPAEpreparedcellslabelledwithMito-TrackerRed,AlexaFluor488phalloidinandDAPItakenbyBillArcher.Aboveler20xmag,aboveright40xmag.
NikonA1-R/MulI-PhotonLaserScanningConfocalMicroscope
TheNikonA1-RConfocalisapowerful,fully-automatedconfocalimagingsystemallowinguserstocapturehigh-qualityconfocalimagesofcellsaswellasmoleculareventsathighspeedswithitsResonantScannerorwithenhancedsensiCvityusingitsSpectralDetecCon.WiththeuseofanenvironmentalchamberyoucanimageinrealCmelivecellsandCssuesthus,makingitidealforabroadrangeofuses.Finally,withthebenefitsthatcomewithtwophotonimaginginMulC-PhotonmodeonecanworkwithliveCssuesfordeepimaging,lowphoto-damagingandhighspeedacquisiConsnotpossibleinnormalconfocalimaging.
ConfocalImagingofRhodopsin:WorkdonebystudentsdoinganUndergraduateResearchcoursetaughtbyDr.MichelleWhaley.TopRightImage:Studentswereimagingrhodopsinproteinintheeyesofmosquitos.SlideswerepreparedwithcareintheHistologyFacility.BoJomRightImage:AlexMetoxen,undergraduateresearcherinDr.J.O’Tousa’slab,wasexaminingrhodopsin(green)migraContolightsensiCvephotoreceptormembranes(red)intheeyeofthefruitfly.
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DeepimagingofliveIssue:TheinfraredlaseremployedinthemulC-photonexcitaConmodalityoftheNikonA1-R/MPisbestsuitedforlong-termlivescanningdeepintotheCssue.Atthelerareimagesofa3DculturedtumortakenbyAmyLeliaertfromthelabofDr.ZachSchafer.Thetwo-photoncapabilityallowsimagingwithmulCplefiltersetsthroughtheenCrecyst,togeneratethreedimensionaldatasetsoftheseclustersoftumorcells.
InaddiIon:Off-lineImageProcessingStaIonwithAutoQuantDeconvoluIonSoqware
(Bright-fieldandConfocal)
…becauseyoushoulddeconvolveeverything!
Device Live-cellenvironmentalchamber?
Approximatesample
penetraIondepth
(micrometers)
BlurminimizaIonmechanism
UniquemodaliIes
NikonAZ100Macroscope
no 1,000+ none 8xzoom
NikonEclipse90i no 1,000+ none -
GEHealthcareDeltaVsion
YES
400
deconvoluIon
image
deconvoluIon
Andor/NikonSpinningDisk
YES
200
spinning-diskconfocality
FRAPPA,TIRF,
FRET
NikonC-2
no
200–300
laser-scanningconfocality
-
NikonA1-R
YES
400–600
laser-scanningconfocality
FRET
NikonA1R-MP
YES
600-800
mulIphotonexcitaIon
infrared-laser-
inducedmulIphotonexcitaIon
Whichdevicewillbestservetheneedsofyourexperiment?
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ServicesincludeCssuefixaCon,processing,embedding,
secConing,andstainingofparaffinorfrozenCssuesecCons.
StainingofCssuesecConsrangesfromrouCneH&E’s
(Hematoxylin-Eosin)tospecialstainsdemonstraCngspecificCssuestructures.
TheCoreoffersimmunohistochemicalstainingwithanCbodiessuppliedbyinvesCgators.
HISTOLOGYCOREAboutUsTheNotreDameHistologyCore(NDHC)providesameanstoexaminebiologicalprocessesinmice,rats,frogs,zebrafish,fruitflies,sheepbone,andevenhumanCssuebyuseofimmunohistochemicaltechniquesandpathology.
INFLAMMATIONHematoxylinandEosininLiver
APOPTOSISCaspase-3inOvary
ANGIOGENESISVEGFinKidney
PROLIFERATINGCELLSPCNAinBone
Services
EquipmentThefacilityislocatedinFreimannLifeSciencesanditisequippedwithaShandonCitadelTissueProcessor,Leicamicrotome,Tissue-TekIIIembeddingstaCon,andanewLeicaCryostatwithaspecialTungstenbladeaJachmentcapableofslicingthroughbone.
SarahChapmanBiologicalImagingAssistant
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FABRICATIONLABORATORYNowopenforbusinessacrosscampus!
AboutUs:Thisfacilitymaintains3DprinCngandlasercuungequipmentforrapidprototypingofpartsforresearch,entrepreneurship,orotherapplicaCons.AvailableforproducConanddesignconsulCng.Locatedin010GalvinHall.
Equipment:
Objet30PrimeUsefulwithhard(Vero)andsor(Tango)seriesliquidresinsfrom
Stratasys
UniversalVLS6.60LaserCu-er/Engraver65WaJlaserwith18”x32”
workingarea
TazFDM3DPrinterFilamentbasedprinter
LowerresoluCon,lowercost
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