May 2016 (Provisional)

EDT/Collect

for DigitalMicrograph

Data Collection for Electron Diffraction Tomography

EDT/Collect Manual 1.0

HREM Research Inc.

2

IntroductionTheEDT/CollectsoftwarehasbeendevelopedbyHREMResearchInc.incollaborationwiththe

AnaliteXcompany(www.analitex.com).

TheEDT/CollectDigitalMicrographpluginallowstheusertoCollectbeam-tiltElectronDiffractionTomography3Ddatasets.

GeneralenquiriesontheEDT/CollectforDigitalMicropraphshouldbesentto:

HREMResearchInc.14-48MatsukazedaiHigashimatsuyamaSaitama355-0055Japan

email: support@hremresearch.comwebsite: www@hremresearch.com

EnquiriesonEDT/Collectofatechnicalnatureshouldbedirectedto:

AnaliteXcompanyemail: support@analitex.comwebsite: www.analitex.com

TheproperreferencetothedatacollectionmethodasofJanuary2013is

M.GemmiandP.Oleynikov.Scanningreciprocalspaceforsolvingunknownstructures:energy

filtereddiffractiontomographyandrotationdiffractiontomographymethods.ZeitschriftfürKristallographie-CrystallineMaterials:Vol.228,No.1,pp.51-58.

CopyrightStatements

©Copyright2016HREMResearchInc.andAnaliteX

Allrightsreserved.Thismanualisprotectedbyinternationalcopyrightlawsandtreaties.Unauthorizedreproductionanddistributionofthismanual,oranyportionofit,willbeprosecutedtothemaximumextentpossibleandmayresultinseverecivilandcriminalpenalties.

PortionsofthisdocumentwerepreparedbyHREMResearchInc.byeditingthematerialssuppliedbyAnaliteX.

DigitalMicrographisatrademarkofGatanInc.

3

Installingtheprogram

EDT/CollectisavailableasaplugintoDigitalMicrographoftheGatanmicroscopySuite(GMS).Asof2016thefollowingGMSversionsaresupported:

• 32bit:GMS1.xandGMS2.x;

• 64bit:GMS2.xandGMS3.x;

EDT/Collectisavailableinthe32bitsversion.

TheEDT/Collectsoftwarecontainsonlyproprietarylibrariessuchas,forexample,Intel®MKL.

SoftwarerequirementsThefollowingisalistofthesoftwarerequirementsnecessarytoruntheEDT/Collectplug-in:

- DigitalMicrograph(GATANTM)- USBKeyDriver- HREMMouseToolPlug-in(Free-waredownloadablefromwww.hremresearch.com)- IPUPlug-in(Free-waredownloadablefromwww.hremresearch.com)- GatanEMControlPlug-in(aplug-insupportedbyGatantocommunicatewithamicroscope)- AnaliteXTEMServer(pleaseconsultthemanualfortheTEMserver)

SoftwareInstallationInstallingUSBKeyDriver

Theuserkeydrivershouldbeinstalledbyfollowingtheinstructionsgivenbythekeydriverinstaller.ThekeydriverinstallercomeswithEDT/Collect,oryoucanfinditonourwebsite.

InstallingEDT/CollectPlug-in

Theplug-incanbeinstalledbydrag-and-dropcopytothefolder“PlugIns”(ThePlugInsfoldershouldexistunderanormalinstallationoftheDigitalMicrograph.)

WhentheDigitalMicrographislaunchedafterplacingtheplug-insintothePlugInsfolder,EDT/Collectmenucommandswillappearunder“EDT/Collect”menu.InstallingMouseTools

AllthefilesrelatingMousetoolplug-incanbeinstalledbydrag-and-dropcopytothefolder“PlugIns.”(ThePlugInsfoldershouldexistunderanormalinstallationoftheDigitalMicrograph.)

WhentheDigitalMicrographislaunchedafterplacingtheplug-insintothePlugInsfolder,theMousetoolwillappearasanadditiontothestandardtools.InstallingIPUPlug-in

EDT/CollectusessomefunctionsbasedontheIntel’MKL(MathKernelLibrary)providedbytheIPUplug-in.AllthefilesrelatingtheIPUplug-incanbeinstalledbydrag-and-dropcopy.PleaseconsulttheReadMefilethatcomeswiththeIPUplug-in.InstallingAnaliteXTEMServer

PleaseconsultthemanualfortheTEMserver.Ifyouneedanyhelptoinstall/usetheTEMServer,pleasecontactwithAnaliteXdirectly.

4

AccessingthepluginfromtheDMmenu

EachmoduleisaccessiblethroughthemainDMmenuEDT/Collectandhasauniquenamethatrepresentsthecontentofthecorrespondingmodule.

Theavailablecommandsthroughthemainmenuare:

Calibration…–theTEMcoilscalibration;

CollectData…–themaindatacollectionmodule;

Settings…–changingtheprogramsettings(forserviceoperators,notrecommendedtouse);

Help–gettinghelpontheplugin.

About…–displaystheAboutdialogwiththeplugininformation.

5

Pluginsettings

AftertheinstallationoftheEDT/CollectpluginintheDigitalMicrographtheusershouldsetupsomeparameterssothatthepluginknowswhatCCDcamerascanbeused,thediffractionpatternscalebars(pixelsize)foraccessiblecameralengths,availablesampleholdersandetc.

InordertoaccessthepluginsettingspleasechoosetheSettings…menuitemfromtheEDT/CollectDMmainmenu.Thefollowingdialogwillshowup:

CCDsettings

ThepluginwillautomaticallyidentifyallcamerasavailabletoDigitalMicrographthatcanbeaccessedthroughtheCurrentcameradropdownlist.

ForeachcameratheusercansettheOversaturationLevel(willbeusedbytheEDT-Processsoftwarepackagetocountoversaturatedpixels).

IMPORTANT:Theusermustaddcalibrationvalues(pixelsize)atleastforasingleTEMcameralengthvaluethatisplannedtouse.ThesevalueswillbeusedbytheEDT-Processprograminordertocalculatecorrectd-spacingvaluesoftherecordeddiffractionpatterns.

Fordiffractionmodecalibration:

1. SelectSADIFFfromtheCalibrations:Mode:dropdownlist(defaultisSAMAG);2. PressAddbuttoninordertoaddacalibrationitemtothelist.(Ifyouneedtoremovethe

selectedcalibrationitem,thenselecttheIDtobedeleted,andpresstheRemovebutton.)

Thefollowingdialog,thatshowsdefault(uninitialized)values,willappear:

6

3. Fillinthevalues:(A)ID–theintegeridentifier(uniqueindex)ofthemagnification/cameralength;

(B)Mag/C.L.–themagnificationvalueorcameralength(in[mm])thatiscurrentlysetattheTEM;(C)Scale–thepixelsize.Ifyoudon’tknowthecorrectscalevalue,escapethedialogby

pressingCancelbutton.Youcanobtainthecorrectscalevalueasexplainedbelow.

Inputexample

4. PressOK.5. Repeatthesteps2-4forallcameralengththatareusedforthedatacollectionindiffraction

mode.

HowtogetacorrectscalevalueInordertogetthecorrectscalevalue(angstrom/pix)theusercanusethescalebaraccessiblefromtherecordedelectrondiffractionpatternusingstandardDMacquisition.

1. SetthedesiredcameralengthonyourTEM.NOTE:onsomemicroscopesthecameralengthchangesdependingifthefluorescent(view)screeninthecolumnisupordown(FEITEMs).IfthisisthecasepleaseputthescreenupsothatyourCCDcanrecordarealdiffractionpattern.

2. Acquireadiffractionpatternatbinning1(important!)andanyexposure(canbeveryshortasthediffractiondataitselfisnotneeded).

3. Afterthepatternacquisitionisdoneright-clickonthepatternwiththemouseandselectImageDisplay...fromthecontextmenu:

7

4. FindandselecttheitemImageàCalibrationontheleft:

5. FindtheScalevalueintherightpartoftheImageDisplayInfodialogunderDimensionsgroup.

NOTE:DMhastwoscalevalues–forx-andy-coordinatesoftheimage(forexample,ifimagebinningalongx-axisisdifferentfromthebinningalongy-axisduringtheacquisition.CurrentlyEDT/Collectsupportsonlyasinglescalefactor.

NOTE:DMusesreciprocalnmunits(nm–1)forthescalefactor.EDT/CollectusesreciprocalÅngströmunits(Å–1).FortheabovescalevaluetakenfromDM(0.0395259[1/nm])usethevalueof0.00395259[1/Å]whenaddingacalibrationitemtothelistintheEDT/CollectCCDsettingsdialog.

8

Sampleholdersettings

ThistabwillhelpyoutomaintainthelistofsampleholdersthatcanbeusedonyourTEM.Theusercanadd/remove(usingAddandRemovebuttonsrespectively)sampleholdersfromthelist:

TheRemovebuttonwillremovecurrentlyselectedsampleholderfromthelist.

WhenpressingtheAddbuttonthefollowingdialogwillappear:

Heretheusercanspecify:

l thenameoftheholder(Nameeditbox);

l thetiltinglimitoftheholderindegrees(alphatilt,Limiteditbox);l theinitialdelay(inmilliseconds)ofthestagebeforeitwillstarttilting(FirstStageDelay).

ThisvalueisneededifordertohavesomedelaybeforetheCCDwillbeactivatedinthe

Viewmodesothattheusercanseethereisthecrystalwillmoveoncethestagewillstarttilting.

l Theconsequentstagedelayvalueisreservedforthefutureuse.

9

Plug-inparameters

TheParameterstabisusedtosetuptheEDT/Collectinternals.

TheTEMModecalibrationgroupcanbeonlysetafterthefirstdatacollectionhasbeenprocessedbytheEDT-PROCESSsoftware.EDT-PROCESSwillreporttheAlphaTiltAxisandtheneedoftheinversion

ofthebeamtiltdirectionand/ortheEwaldsphere.

TheCollectDatagroupshouldbeusedonlyforthecameraswithaphosphorscreen(suchasGatanErlangshenES500W)inordertointroducesomedelaybetweentheconsequentacquisitionofdiffractionframesgettingridofthepreviousdecayingdiffractionpatternthatmighthavestrong

intensitiesthatcouldaffectthecurrentframe.Inallothercasesitissafetoremovebothcheck-boxmarks.

10

TEMcoilscalibrationSinceonsomeelectronmicroscopes(forexampleJEOL)thechangesofcoilsandlensesaredoneininternalunitsthecalibrationofthesecoilsisneededinorderto:

• Convertthearbitraryunitsintophysicalunits(e.g.nmordegrees);

• Compensatethebeamshiftinducesbyonelensusingtheother.

Thecalibrationofeachcoilisdonein2dimensions:alongX-andY-axesofthecorrespondingcoil.ThecalibrationisperformedautomaticallybytheprogramafterpressingthecorrespondingCalibrate

button.

ThecalibrationmustbeperformedintheDIFFRACTIONmodeofthemicroscope.FortheTEMmodeoftheJEOLmicroscopeinserttheSAEDapertureandfocusthebeamintothepoint.FortheNBD

modeselectthesizeandthebrightnessofthebeam(sameaswillbeusedlaterforthedatacollection)andfocusthebeamintothepoint.

PleaseselecttheCalibrationfromtheEDT/CollectmenuinordertocalibratecurrentTEMcoils.Thefollowingdialogwillbeshown:

Forthe3DEDTdatacollectionpleaseusethefollowingsteps:

1. Selectanemptyareaonyourgrid;2. InsertasmallestSAEDapertureaccessibleonyourmicroscope;3. Spreadthebeam;

4. SwitchintodiffractionmodeonyourTEM;5. Setthedesiredcameralength;6. Centerandfocusthecentralspot;

7. ChecktheRecalculateangleontheCalibrationdialog;8. SetsomevalueinChangevalue:editboxfortheBeamtiltcoils.AninitialvalueforJEOLTEM

canbe1000unitsforthestart;

Beamtilt

Projectorcoil

11

9. PressCalibratebuttonandwaitwhilethecalibrationisfinished;

10. Repeatsteps8–9fortheProjectorcoil.

CAUTION!BecarefulwiththedirectexposureoftheCCDtotheelectronbeam!Useminimumintensitybeam(spreadthebeamusingtheBrightnessknoboftheJEOLTEM;usehighspotsize,forexample5;insertsmallSAEDapertureinTEMmode).

Thecalibrationoftheselectedcoilisdoneautomaticallyaccordingtothefollowingsteps,whenyoupressCalibratebutton.Thecalibrationprogram

1. readstheuser-suppliedChangevaluefromthepanel(let’scallitCVvalue);

2. readscurrentcoilvaluesforx-andy-axes(let’scallthemasX0andY0values);3. recordsthediffractionpattern(originalpattern);4. changesthex-axisvalueofthecoilbytheCVvalue(soitbecomesX0+CV);

5. recordsthediffractionpattern(X-modifiedpattern);6. changesthex-axisbacktotheoriginalX0valueandchangesy-axisvalueofthecoilbythe

CVvalue(soitbecomesY0+CV);

7. recordsthediffractionpattern(Y-modifiedpattern);8. restoresthecoily-axisvaluebacktoY0;9. calculatestheshiftsofthespotusingnormalizedcross-correlation;

10. iftheshiftsissmallandthecheckboxforAutomaticChangevalueisselected,theCVvalueisautomaticallydoubledandgotothestep4;

11. savesthecalibrationmatrix(thedirectionofthecoilx-andy-axesandthemeasureddeviationsalongx-andy-axes).

Thecorrectcoilcalibrationimagesshouldlooksimilartothefollowing:

Current Shifted Cross-correlation

TheCurrentpatternisasumof3recordeddiffractionpatterns–theoriginaland2withinducedchangesalongx-andy-axesofthecoil.The3spotsseenontheCurrentpatterncanbeinterpretedasshownontheFigurebelow:

12

NOTE:thecalibrationwillbemorepreciseifthe2spotsobtainedfromtheinducedchangesofx-andy-axesofthecoilarewillseparated(closetotheedgeoftheCCDframe).ThesepositionsdependontheChangevalueforthecorrespondinglens.

TheShiftedpatternshowsthelastpatternwiththedeviationofthey-axisofthecoil.

TheCross-correlationpatternisthesumof2individualcross-correlationpatterns:(1)theoriginalpatternandthepatternwherethex-axishasbeenchanged;(2)theoriginalpatternandthepatternwherethey-axishasbeenchanged.Sinceineachcasethecross-correlationshowsasinglepeakthen

thesumofthe2cross-correlationfunctionsmustshow2well-separatedpeaks.

TheoriginalspotThespotafterx-axischange

Thespotaftery-axischange

X

Y

13

Collecting3DEDTdatasets

NOTE:ItisstronglyrecommendedtoruntheTEMcoilsCalibrationmodulepriorthedatacollection.

Thedatacollectionof3DElectronDiffractionTomographydatacanbeperformedusingthe“CollectData…”modulefromthemain“EDT/Collect”menu.TheDataCollectiondialogboxwillbeshownasitcanbeseenonthefollowingfigure:

Thegroupbox“SampleHolder”canbeusedtoselecttheappropriatesampleholderthatiscurrently

inuse(seesection“Sampleholdersettings”onhowtomanagealistofsampleholders).Itisrequiredtochoosethesampleholderpriorthedatacollection.Theactuallimitofthealpha-tiltangleswillbeshownbelowtheselectedsampleholdertype(“Limit(deg):70”onthefigureabove).

Thegroupbox“CollectData”shouldbeusedforthedatacollection.

Thesettingsthatwillbeusedforthedatacollectionare:

l CollectiontypecanbechosenfromtheCollectdrop-downbox(currently“Rotation3D

(manual)”);l Thefolderwheretheacquiredframeswillbestoredcanbechosenusingthebrowsebutton

(“Folder…”).Theselectedfolderwillbeshownexttothebutton,forexample,

(“C:\Users\VALUEDGATANCUSTOMER\Desktop\Test\”onthefigure);l TheBeamtiltrange(indegrees)correspondstothefullanglethatwillbecoveredbythe

beamtiltofTEM.Incaseof3°thebeamtiltwillvaryfrom–1.5°to+1.5°.Inthisexamplethebeamtiltwillcovertheanglesfrom–0.5°to+0.5°(1°intotal);

14

l TheBeamtiltstep(indegrees)specifiesthebeamtiltstepthatwillbeusedinordertocovertherangeofthebeamtiltanglesspecifiedinBeamtiltrange.Inthisexampletheprogramwillchangethebeamtiltintherangefrom–0.5°to+0.5°(covering1°)withastepof0.5°.Thismeansthatthebeamtiltwillbeconsequentlysettothevaluesof–0.5°,0°and+0.5°(3frames).IfwesettheBeamtiltrangeto3°andkeepthestepof0.5°thenthebeamtiltwillbeconsequentlysettothevaluesof–1.5°,–1°,–0.5°,0°,+0.5°,+1°and+1.5°(7frames);

l TheSeriesoverlap(indegrees)definestheoverlapbetweenconsequentgoniometertilts.Thisvalueisinordertocompensatethemechanicalimperfectionsofthestage.Inidealcasethegoniometershouldtiltbythesamevalueasthe“Beamtiltrange”(1°inthiscase).However,ifthevalueof“Seriesoverlap”isdifferentfrom0thentherealgoniometeralpha-tiltwillbe“Beamtiltrange”minus“Seriesoverlap”(1°–0.1°=0.9°inourexample).The“Seriesoverlap”valuecanbekeptas0.2°formostdatacollections;

l TheStartdelay(milliseconds)andtheBeamblankcheckboxareusedforsometypesofCCDsthataresensitiveforthebeampositionchanges.SomeCCDcameras(suchasGatanES500W)useafluorescentscreensothattherecanbeatraceleftbythestrongbeamwhileitmovesduringthebeamtiltorshift.ItmaydecayslowlyandcanbenoticedontheCCD.InordertocompensatefortheseeffectspleasechecktheBeamblankandputsomedelay(500-1000msshouldbegenerallyenough).

NOTE:Thecollecteddatasetmustoccupyanindividualfolder.Everynewdatacollectionmustbepointedintoanewfolderotherwiseallpreviousdatawillbeoverwrittenwiththenewdataset!

15

Datacollectionstep-by-step

InordertostartthedatacollectiontheusedneedstopresstheStartbuttonatthebottomofthepanel.

Thedatacollectioncontainsthefollowingsteps:

1. Checkthesampleheight(usingtheTestFocusorTestWobblebuttons);

2. Collectthebeamtiltdata(usingtheCollectDatabutton);3. Tiltthestage(usingtheTiltstagebutton);4. ManuallytrackthecrystalusingtheCCDcamera;

5. Repeatfromthestep1;6. FinishthedatacollectionbypressingtheCancel(Finish)button.

CheckingthesampleheightTherearetwowaystocheckthecrystalheight(inimagingmode!):(i)thedynamicwobbleofthebeambytiltingand(ii)subtracttwoframesrecorderwithoppositebeamtilts.

ThedynamicwobbletestissimilartotheJEOLX-WOBBLEbutton.Inthiscasethedatacollection

programinduces2oppositebeamtilts(+/–0.5°inourcase)andswitchestheCCDintotheviewmodefortheobservations.Thecorrectsampleheightwillbetheonewhenthecrystalrelativemovementwillbeminimal.

Theframesubtractionproducesthefollowingimagesinthecaseof(left)thecorrectsampleheight–

noticetheaveragegrayleveloftheinnerpartoftheSAEDaperturewithminimumnumberofdistinguishablefeaturesand(right)thesampleheightiswrong–thereare2shadowsofthecrystalcorner.

Twoshadowsofthecrystalduetothesamplemovementcausedbythebeamtilt

16

CollectingthedataThebeamtiltdatawillbecollectedautomaticallybytheprogram.Alltheframeswillbesavedtodisk

intheDM3/DM4format.Thetypicalstatusofthedatacollectionisshownonthefigurebelow:

NOTE:Duringthedatacollectionofthebeamtiltseriesallbuttons(exceptCancel)willbedisabled.

TiltingthestageAsitwasmentionedpreviously,thestagewillbetiltedbytheanglethatiscalculatedastheBeamtiltangleminustheSeriesoverlapangle.Inthecasethatispresentedinthismanualthestagetiltwillbeequivalentto0.9°(1°–0.1°=0.9°).

ThedatacollectionprogramwillswitchtheCCDintotheviewmodeforthecrystaltracking.

ManuallytrackingthecrystalThisisthemostcrucialstepduringthedatacollection.Theusermustmovethecrystalbacksoitwilloccupytheappropriatepositionsothatthenextbeamtiltseriescanbecollected.ThecrystaltrackingcanbeeasilyachievedwiththeX-Ystagecontrolofthemicroscopeafterthestagehasbeen

tilted.

17

DataprocessingAlltherecordedframesandthedatacollectionlogfilewillbestoredintheselectedfolderforthecurrentdatacollection.Donotchange/erase/modifythefilesinthefolder!Thewholefolderhowevercanbemovedtoanyotherlocation.

ThecollecteddatasetcanbefurtherprocessedwiththedataprocessingprogramEDT-PROCESSfrom

AnaliteX.

Bugreporting

Therearelogfilescreatedbytheprogramduringeveryrun.TheselogsarestoredinaspecialfolderthatdependsontheWindowsversion.