fall 2016 newsle-er - notre dame integrated imaging facility · fall 2016 newsle-er ... in vivo...

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,DirectorofNDIIFsmith.115@nd.edu

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:sergei.rouvimov.1@nd.eduTel: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

wleevy@nd.edu

SarahChapmanAssistantDirectorofBiologicalImaging

sarah.chapman55@gmail.com

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(archer.2@nd.edu).

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

Directorsarah.chapman55@gmail.com

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FABRICATIONLABORATORYNowopenforbusinessacrosscampus!

AboutUs:Thisfacilitymaintains3DprinCngandlasercuungequipmentforrapidprototypingofpartsforresearch,entrepreneurship,orotherapplicaCons.AvailableforproducConanddesignconsulCng.Locatedin010GalvinHall.

TonyVanAvermaeteFabLabCoreManagertonyvan22@gmail.com

Equipment:

Objet30PrimeUsefulwithhard(Vero)andsor(Tango)seriesliquidresinsfrom

Stratasys

UniversalVLS6.60LaserCu-er/Engraver65WaJlaserwith18”x32”

workingarea

TazFDM3DPrinterFilamentbasedprinter

LowerresoluCon,lowercost

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