Zeiss AxioObserver D1 Manual de usuario

ZeissAxioObserver.D1Microscope



Room:3164A



StartUpProcedure



1. Removedustcoverfrommicroscope

2. ToggleFluoArcLampswitchON

3. TogglePowerBarswitchON

4. TurnMicroscopeON

‐bypressingpowerbuttonlocatedonleftsideofmicroscope

5. TurnONcomputer

6. Logonto:ObserverUseraccount(nopassword)

7. OpenProgram:AxioVisionRel.4.8



ShutDownProcedure



1. Ifusedoil,gentlyremoveoilfromobjectiveusingneatlyfolded

drylenspaper

2. Put10xobjectiveinplace,andlowerobjective

3. CloseAxioVisionprogram

4. Shutdowncomputer.Waituntilcompletelyturnedoff

5. TurnMicroscopeOFF

6. TogglePowerBarswitchOFF

7. FluoArcLamp:

‐Regularworkhours:setintensityto50%oncontrolpad(F2),

andleavelampON

‐Non‐workhours:TurnlampOFF

8. Covermicroscopewithdustcover,butDON’TCOVERTHEHOT

LAMPHOUSING!!



Microscope Quick Guide



ZeissAxioObserver.D1MicroscopeGuide

Room:3164A

StartUpProcedure

1. Removedustcoverfrommicroscope

2ToggleFluoArcLampswitchON

3TogglePowerBarswitchON

4TurnMicroscopeON

‐bypressingpowerbuttonlocatedonleftsideofmicroscope

5TurnONcomputer

6Logonto:ObserverUseraccount(nopassword)

7OpenProgram:AxioVisionRel.4.8



ConfiguringImagingSetup

1. Ifnotalreadyvisible,opentheWorkareawindow

2. FromtheWorkareamenu,double‐clickonMultidimensionalAcquisitiontoexpandlist,andthen

selectMultidimensionalAcquisition.

3. Underthetab,clickLoadbutton.Selecttheappropriateexperimentset‐updependingon

whatchannelsyouwanttoimage(ieAllChannels).

4. Selectthecolorstab

5. Youcanactivate/deactivatechannelsbyright‐mouseclickingonthechannelnumber



SetSampleonStageandFocus

Onlyuseslidesthathavebeenthoroughlycleanedofmountingmedium!

1. Choosetheappropriateobjectiveonmicroscope

2. Lightlycleanyourslide(onbothsides)withakimwipeandasmallamountof70%ethanol.

3. Ifusinganoillens,addaverysmalldropofimmersionoil.

4. Loadslideonstage.

5. Bringspecimenintofocusandbringyourareaofinteresttothecenterofthefieldofview.



CapturinganImage

1. Underthetab,selectyourchannelandclicktheMeasurebutton.Alivepreviewwindowappears.

2. Makesurethe‘Lineardisplayoffullrangeofpossiblevalues’buttonisselected.



3. Adjusttheexposuretimetogetanappropriatesignalintensity.ClickOKbuttononleftside.



4. Repeatexposuretimemeasurementforeachchannel,however,adjustthefinefocusforonlyone

channeliethemostimportantone

5. Tocaptureyourfinalimage,pressthebuttonlocatedatthebottomoftheCtab.



Post‐AcquisitionAdjustments

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Brightness/Contrast

1. Temporarilyturnoffthepseudo‐coloringofthedisplayedimagebypressingthebutton.Each

channelisdisplayedinblack/white;youcanviewonechannelatatime.

2. Clickonthepropertiesbuttonfoundbelowtheimage



3. WhileunderDisplaytabofpropertieswindow,clicktheMin/Maxbutton,orunderyourimage,press

thebutton.

4. Youcanmakefurtheradjustmentsusingthedisplayedhistogram(iemovetopoflineleft,andbottom

oflineright)

‐NOTE:donotadjusttheGamma

5. Repeatforeachchannel

6. Pressthebutton



AddingaScalebar

1. Toaddascalebar,pressthebutton,orAnnotationsfromthemainmenuandselectScalebar.

2. Dragcursoralongimagetogetdesiredscalebarsize



SavingFiles

1. Fromthemainmenu,selectFileSaveas

2. SaveyourfilesintheAxioVisionformat(.zvi)intheappropriatedestinationyourlabfolderyour

folder.PleaserefertotheAttentionUserssignspostedonthewall.



ShutDownProcedure

8Ifusedoil,gentlyremoveoilfromobjectiveusingneatlyfoldeddrylenspaper

9Put10xobjectiveinplace,andlowerobjective

10 CloseAxioVisionprogram.Donotsavechangesmadetotheexperiment!

11 Shutdowncomputer.Waituntilcompletelyturnedoff

12 TurnMicroscopeOFF

13 TogglePowerBarswitchOFF

14 FluoArcLamp:

‐Regularworkhours:setintensityto50%oncontrolpad(F2),andleavelampON

‐Non‐workhours:TurnlampOFF

15 Covermicroscopewithdustcover,butDON’TCOVERTHEHOTLAMPHOUSING!!



MicroscopeControls

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DisplayScreen



TransmittedLightControls



ReflectedLightControls



LightPath

Locatedonleftsideofmicroscope



 



ObjectiveType

ObjectiveNA

ObjectiveContrastMethod

ObjectiveMagnification

CondenserContrastMethod

HALLampVoltage

TLShutter(open)

RLShutter(closed)

Reflector

Locatedonbottomfrontof

microscope

ControlsHALlamp

voltage/intensity

Dia

hra

mControl

Condensercontrast

method

Condensera

erturecontrol

Locatedonbottomright

ofmicroscope

Opens/closesFLshutter

TurnsHALLampon/off

Locatedonbottomright

ofmicroscope

Locatedatbackofright

focusknob

Movesreflectorturret

50%Camera

50%Ocular

100%Camera 100%Ocular

Objectives

 

ZeissAxioObserver.D1MicroscopeObjectives

Room:3164A

 123

 A‐Plan10xLDPlan‐NeoFluar20xPlan‐Apochromat

20x

0.25Ph10.4KorrPh2‐M270.8M27

Magnification10x20x20x

NumericalAperture0.250.40.8

WorkingDistance(mm)4.27.9atcoverglass0.750.55

Coverglass(mm)0.170‐1.50.17

ImmersionNoneNoneNone

LongDistance(LD)‐ Yes‐

CorrectionRing(Korr)‐ Yes‐

OpticalSystemICSICSICS

Flatness***********

ColorCorrection***********

UVTransmission********

IRTransmission*********

DIC‐ ‐ (DICII)*****

PhaseContrastPh1Ph2‐

Catalog#441031‐0000‐000421351‐9970‐710420650‐9901‐00

 456

 Plan‐NeoFluar

40xPlan‐Apochromat63xPlan‐NeoFluar5x

 1.3OilPh31.4OilDICM270.15

Magnification40x63x5x

NumericalAperture1.31.40.15

WorkingDistance(mm)0.20.1913.6

Coverglass(mm)0.170.170.17

ImmersionOilOilair

OpticalSystemICSICSICS

Flatness************

ColorCorrection************

UVTransmission***********

IRTransmission**********

DIC(notequipped)**(DICIII)*****‐

HighContrastDIC‐ Yes‐

PhaseContrastPh3‐ ‐

Catalog#000000‐1022‐819420782‐9900‐000440320‐0000

ICS=InfinityColorCorrrectedSystem



Plan‐NeoFluar100x

1.3OilPh3

100x

1.3

0.2

0.17

Oil

ICS

****

***

‐

Ph3

000000‐1031‐171



Filter Sets



ZeissAxioObserverD1MicroscopeFilters

Room3164A



1. TransmittedLight



2. FilterSet34(DAPI/BFP)

Excitation:BP390/22

BeamSplitter:FT420

Emission:BP460/50



3. FilterSet38(Cy2/GFP)

Excitation:BP470/40

BeamSplitter:FT495

Emission:BP525/50



4. FilterSet46HE(YFP)

Excitation:BP500/25

BeamSplitter:FT515

Emission:BP535/30



5. FilterSet45(Cy3.5/mcherry)

Excitation:BP560/40

BeamSplitter:FT585

Emission:BP630/75



6. FilterSet50(Cy5)

Excitation:BP640/30

BeamSplitter:FT660

Emission:BP690/50

















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