Carl Zeiss Axio Imager microscope User manual
Below you will find brief information for microscope Axio Imager. The Axio Imager is a research-grade microscope that offers a wide range of features and capabilities. You can set up transmitted-light brightfield, darkfield, phase contrast, and differential interference contrast (DIC) microscopy. The microscope can be configured for biological or materials applications. You can even adjust the brightness of the TFT display and configure the assignment of functions for the various buttons on the microscope. This manual will help guide you through the operation of the Axio Imager.
Advertisement
Advertisement
OPERATION
Axio Imager Operating the motorized microscope via touchscreen of TFT display Carl Zeiss
•
To start configuring a nosepiece position, press the corresponding button.
In the Configure Objective # popup window, you can choose among various options:
− Manual button
You have to enter magnification, NA and immersion type on your own.
− From list button
Select the magnification from the Preselect
Magnification list and the appropriate objective from the Objective list.
− Via Cat. No. button
To select the objective, enter the Zeiss Cat.
No. (XXXXXX-XXXX-XXX).
Fig. 4-37 Microscope -> Settings ->
Components -> Objectives
You can delete an existing objective configuration by pressing the Empty Pos. button. For that, choose the corresponding nosepiece position and confirm the safety query with Yes.
•
•
Press the Save button to save the objective configuration for the selected nosepiece position or close the popup window without saving the configuration by means of the Cancel button.
If you are about to overwrite an existing objective configuration, answer the safety query with Yes.
When entering the 15-digit Zeiss Catalogue No., the leading six zeros or the following seven zeros (after 123456 enter a hyphen (-) or enter 1234-567 and press OK). The missing zeros will be added automatically.
(2) Reflector
With the controls on this tab, you can configure the reflector turret.
The tab shows up to 10 buttons dependent on the actual number of positions on the reflector turret, which is detected by the system during initialization (and on activation of the
Settings/Components page). As long as you have not configured any reflectors yet, the buttons are labeled only with the numbers of the turret positions.
After you configured a reflector position, the following data will be displayed: Designation
(Type), Reflected-light module (RL), Transmittedlight position / module (TL)
Fig. 4-38 Microscope -> Settings ->
Components -> Reflector
After you configured a reflector position, the corresponding reflector button on the
Microscope/Operation page is labeled accordingly.
B 46-0046 e 12/05 101
OPERATION
Carl Zeiss Operating the motorized microscope via touchscreen of TFT display Axio Imager
Fig. 4-39 Microscope -> Settings ->
Components -> Reflector
•
•
•
•
To start configuring a reflector turret position, press the corresponding button.
In the Configure reflector position # in
reflector turret popup window, you can choose the respective reflector from the list. In the Resulting Configuration line, you can read the current selection.
Press the button for RL and/or TL.
Press the Save button. If the turret position was already configured, a safety query will appear.
(3) Focus
On this tab, you can enter the firmware settings for the focusing drive. The speed of the focusing drive is adjustable individually for every objective.
Up to seven buttons are displayed dependent on the actual number of positions on the nosepiece, which is determined by the system during initialization (and on activation of the
Settings/Components page). As long as you have not configured any objectives yet, the buttons are labeled only with the numbers of the nosepiece positions.
After you assigned an objective to a specific button, the magnification appears on the left, blue half of the button. The right, gray half of the button is labeled with the focusing speed.
Fig. 4-40 Microscope -> Settings ->
Components -> Focus
•
•
If you want to change the focusing speed for an objective, press the respective gray part of the button.
In the Focus Speed for Objective # popup window, set the desired speed using the
◄► buttons.
The higher the numerical value, the higher is the set speed.
•
Press the Save button.
The stage-lowering function is activated or deactivated by means of the ON / OFF buttons.
102 B 46-0046 e 12/05
OPERATION
Axio Imager Operating the motorized microscope via touchscreen of TFT display Carl Zeiss
c) Parfocality
The parfocality function is activated or deactivated by means of the ON / OFF buttons.
The parfocality can be configured via the
Adjustment button. On pressing this button, a wizard appears guiding you through the configuration procedure.
All objectives must be focused successively. Start with all dry objectives from the highest to the lowest magnification. Then, proceed with all immersion objectives from the highest to the lowest magnification. By pressing the Next
Objective button, the nosepiece rotates to the next objective. After all objectives were focused, press the End button.
Fig. 4-41 Microscope -> Settings ->
Components -> Focus
This tab allows you to configure the adapters and beam splitters (beam-splitting mirrors / reflecting mirrors) for the camera ports (Top Port / Back Port / Deflection Plane).
Before attaching/removing the camera path deflection, left (Deflection Plane) or before any tube change, switch off the microscope to ensure that the system detects the correct port status during the next initialization process.
a) Adapter
Up to three buttons are displayed here dependent on the equipment of the camera path deflection and the tube used. The status of the ports is detected by the system during initialization (and on activation of the Settings/Components page).
•
To assign an adapter to a button, press the gray button.
This will bring up the Select Camera Adapter list.
•
Choose the installed adapter from the list using the
▲ ▼ buttons.
•
Press the Save button, to assign the selected adapter to the port. Press the Cancel button to close the window without any selection.
Fig. 4-42 Microscope -> Settings ->
Components -> Camera Ports
The button is now labeled with the magnification factor. Proceed analogously for the other ports.
b) Splitter
Up to three buttons are displayed here dependent on the equipment of the camera path deflection and the tube used. The status of the ports is detected by the system during initialization (and on activation of the Settings/Components page).
•
To select a splitting ratio, press the gray button.
This will bring up the Select Splitting Ratio list.
B 46-0046 e 12/05 103
OPERATION
Carl Zeiss Operating the motorized microscope via touchscreen of TFT display Axio Imager
•
•
Press the buttons for the desired splitting ratio; multiple selections are possible.
Press the Save button, to store the selected splitting ratio. Press the Cancel button to close the window without any selection.
The button is now labeled with the splitting ratio. Proceed analogously for the other ports.
The configured splitting ratios will be offered then for selection on the Light Path tab of the
Microscope/Operation page.
Fig. 4-43 Microscope -> Settings ->
Components -> Stage
(5) Stage
This tab will be displayed if a motorized microscope stage (only CAN bus types) is used. On this tab, the operator can activate or deactivate the
XY motion of the stage for the load / operating position. This has an effect on the function of the
Load Position button on the Microscope /
Control page.
(6) Misc(ellaneous)
On this tab, you can configure further optional microscope components.
The number of buttons displayed depends on the components found during initialization or on activation of the Settings/Components page.
You can customize up to six buttons with the following components:
− Man. condenser
Fig. 4-44 Microscope -> Settings ->
Components -> Misc(ellaneous)
If you use a manual condenser, but a motorized luminous-field diaphragm, you should enter the condenser aperture here.
This is necessary, as the Light Manager automatically adapts the luminous-field diaphragm to the field of view of the eyepieces, and the various condensers have different magnification factors.
104 B 46-0046 e 12/05
OPERATION
Axio Imager Operating the motorized microscope via touchscreen of TFT display Carl Zeiss
−
Illumination
Here you can choose whether you work with a halogen or the LED illuminator.
−
Tube lens turret settings
From the Tube lens turret settings list, choose the magnification factors that apply to the positions 2, 3 and 4 of the turret.
− Field of view of eyepieces
Here, enter the field-of-view number of the eyepieces used. This is necessary, as the Light Manager automatically adapts the luminous-field diaphragm to the field of view of the eyepieces, but cannot automatically recognize the field of view.
−
Camera adapter
Here, enter the magnification factor of the used camera adapter. On the Home page, the selected value will be taken into account when computing the total magnification of the microscope.
B 46-0046 e 12/05 105
OPERATION
Carl Zeiss Operating the motorized microscope via touchscreen of TFT display Axio Imager
− Allocation C-DIC turr. (modulator turret) only for Axio Imager MAT
If a C-DIC turret is used and an Axio Imager MAT microscope is available, choose the appropriate filters for turret positions 2, 3 and 4 here.
−
Selection of reflected-light components
If you use a motorized stop slider, attenuator or a double filter wheel in reflected light, you must specify this component here. The selected settings will be stored by restarting the instrument.
4.8.5.2 User
The User page is accessible by pressing the User button in the navigation button bar. The page contains the following four tabs: Mode, Buttons
Left, Buttons Right and Stand Type.
(1) Mode
On this tab, you can choose between Standard and Personal Settings.
Fig. 4-45 Microscope -> Settings -> User ->
Mode
In Standard mode, all factory-set functions
(delivery state) are active. In the Personal Settings mode, those settings are active that have been defined by an administrator for the following control elements:
− Five buttons on Z focusing drive, right side/ left side
−
Two buttons on base of stand (operating position / load position) right side / left side
Any change of the assignment of the button functions, requires the entry of an administrator password.
Mind to whom you make the administrator password accessible. Arbitrarily changed assignment of the button functions may result in damage to the microscope and specimens!
The factory-set password is "12345".
106 B 46-0046 e 12/05
OPERATION
Axio Imager Operating the motorized microscope via touchscreen of TFT display Carl Zeiss
Any change of the assignment of the button functions, requires the entry of an administrator password. Otherwise, you can only view the assignment, but not edit it.
•
•
On this tab, you can configure the assignment of functions for the Z focusing drive buttons and the buttons on the base of the stand. The control elements are displayed symbolically. The two upper buttons and the bottom buttons on the Z focusing drive are to be configured in pairs.
•
Press the gray button to open the selection list.
Use the
▲ ▼ buttons to choose the appropriate function from the list. Only those functions are listed that are really accessible on the microscope.
Press the Save button to assign the desired function. With Cancel, you can close the window without choosing a function.
Proceed analogously for the other button functions.
Any change of the assignment of the button functions, requires the entry of an administrator password. Otherwise, you can only view the assignment, but not edit it.
To configure the button functions, refer to the description “(2) Buttons Left” above.
On this tab, you can choose whether the
Axio Imager shall behave like a biological/medical microscope or a materials microscope. The changed setting will become active after the automatic restart of the instrument.
(5) Language
On this tab, you can choose the language of the user interface of the TFT display. At present, it is available in English and German. The changed setting will become active after the automatic restart of the instrument.
Fig. 4-46 Microscope -> Settings -> User ->
Stand type
B 46-0046 e 12/05 107
OPERATION
Carl Zeiss Operating the motorized microscope via touchscreen of TFT display Axio Imager
4.8.5.3 Extras
The Settings/Extras page contains five tabs:
Light Manager, Oil Stop, Dazzle Protect(ion),
Ethernet and Misc(ellaneous).
Fig. 4-47 Microscope -> Settings -> Extras ->
Light Manager
On this tab, you can activate and deactivate the
Light Manager. The Light Manager serves for automatic light-intensity adjustment (see 4.7 on how to use it).
Fig. 4-48 Microscope -> Settings -> Extras ->
Oil Stop
On this tab, you can activate or deactivate the Oil
Stop function. This function prevents a dry objective from being swiveled into immersion liquid by lowering the stage whenever you switch from an immersion to a dry objective.
Fig. 4-49 Microscope -> Settings -> Extras ->
Dazzle Protect(ion)
108
Note:
When you deactivate the Dazzle Protection function globally, all other options on this tab appear grayed out.
If your microscope has a Bino Shutter and the
Dazzle Protection function is activated, the other two options (transmitted light / reflected light dazzle protection) are inactive, i.e. the dazzle protection function for these components is deactivated.
If one of the above components is not installed, the corresponding buttons are not available.
B 46-0046 e 12/05
OPERATION
Axio Imager Operating the motorized microscope via touchscreen of TFT display Carl Zeiss
(4) Ethernet
On this tab, you can choose settings for the connection of the Axio Imager via Ethernet.
(5) Misc(ellaneous)
This tab allows you to calibrate the TFT display.
4.8.5.4 Info
The Settings / Info page only contains the Firmware tab.
Fig. 4-50 Microscope -> Settings -> Extras ->
Ethernet
Fig. 4-51 Microscope -> Settings -> Extras ->
Misc(ellaneous)
Fig. 4-52 Microscope -> Settings -> Info->
Firmware
B 46-0046 e 12/05 109
OPERATION
Carl Zeiss Operating the motorized microscope via touchscreen of TFT display Axio Imager
4.8.6 Display
The Display page is accessible from the Home page by pressing the Display button on the navigation button bar. To darken the TFT display, press this button for more than a second. To switch it on again, touch it somewhere.
On the Display page, you can adjust the brightness of the TFT display by means of the
◄► buttons.
You can switch off the TFT display by pressing the
Display off button. To switch the TFT display on again, press this button once more.
Fig. 4-53 Home-> Display
After switching the TFT display off, the display returns from the Display page to the page from which it was activated. This page will be shown after switching the display on again.
110 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9
4.9.1
Illumination and contrast methods
Setting transmitted-light brightfield according to KÖHLER
(1) Application
Transmitted-light brightfield microscopy is the most common of all optical microscopic techniques, as it permits high-contrast or stained specimens (e.g. blood smears) to be viewed easily and quickly.
Beside the so-called direct bundles of rays, the indirect, i.e. the bundles of rays diffracted and scattered at specimen details, are of major importance for imaging true to the object. The higher the portion of indirect bundles of rays (aperture), the more realistic is the microscopic image according to ABBE.
To fully exploit the optical performance of the microscope, particularly that of the objective, condenser, luminous-field diaphragm and aperture diaphragm should be set according to the rules of the KÖHLER illumination principle. These fundamental rules of microscope adjustment are described in detail below in
Section 4.9.1 (3) "Transmitted-light brightfield according to KÖHLER" for the Axio Imager.
− The equipment of all Axio Imager microscopes allows transmitted-light brightfield microscopy.
− For the use of the achromatic-aplanatic universal condenser 0.9 H/0.8-0.9 DF, refer to Section
(3) Setting transmitted-light brightfield according to KÖHLER
•
•
− The microscope has been started up correctly as described in Section 3.
− The microscope has been switched on.
•
Set the toggle switch for the halogen illuminators on the rear side of the instrument to transmitted light.
•
Place a high-contrast specimen on the mechanical stage.
Swivel in front lens of condenser (for objectives
≥ 10x) and use the vertical control of the condenser
the specimen is not touched by the condenser (for setting the stop of the vertical condenser drive,
•
•
•
Turn the vertical control of the condenser drive (4-54/5 or 4-55/3) to lower the condenser until the
edge of the luminous-field diaphragm appears in focus (4-54/B).
B 46-0046 e 12/05 111
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Fig. 4-54 Microscope settings on Axio Imager for transmitted-light brightfield
•
Use both centering screws (4-54/6 bzw. 4-55/1) on the condenser carrier to center the luminous-field
When changing the condenser, normally the luminous-field diaphragm remains centered, unless the centering screws were turned.
•
•
112
To adjust the aperture diaphragm (contrast), remove one eyepiece from the tube and look through the
setting provides optimum contrast at almost ideal resolution, and is therefore the best compromise for the human eye.
Insert the eyepiece back in the tube socket.
B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
Specimen field size and objective aperture change after every objective change as possibly does the centering slightly. Therefore, repeat the adjustment of luminous-field diaphragm and aperture diaphragm to obtain optimum results.
For < 10x objectives, the front lens of the universal condenser (if used) must be folded out and the aperture diaphragm fully opened. In the case of such large fields, the luminous-field diaphragm can also be used for better contrasting by its opening being reduced by a certain range. Closing it too much should be avoided in order not to worsen the evenness of the field of view illumination.
•
•
(4) Setting the height stop on the condenser
•
Loosen the fastening screw of the height stop
(4-55/2) using an SW 3 ball-headed screwdriver.
Use focusing drive to focus on the specimen.
Close luminous-field diaphragm and focus it by
turning the vertical control (4-55/3) of the
condenser.
•
•
Carefully raise the condenser slightly without lifting the specimen.
Tighten fastening screw (4-55/2) of height stop.
Fig. 4-55 Setting the height stop on the condenser carrier
B 46-0046 e 12/05 113
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
4.9.2 Setting transmitted-light darkfield
Due to their transparency, unstained biological specimens, such as bacteria or living cell cultures, are often barely or not at all visible in transmitted-light bright field. This changes markedly if such specimens are viewed in transmitted-light dark field, where the specimen is illuminated using an aperture that is larger than that of the objective used.
In dark field, only the diffracted and scattered light components, which are important for imaging, reach the objective, while the direct unchanged light bundles are directed past the objective. This is one of the reasons why even fine structures can be resolved, which are sometimes below the resolving power of the light microscope and which appear very bright on a dark background.
− Universal condenser with dark-field stop in position D
− Achromatic-aplanatic universal condenser 0.9 H/0.8-0.9 DF (424216-0000-000), for low magnifications with wide-field-DF-slider for 2.5x-5x (424215-0000-000)
− Use of ICS objectives up to the maximum aperture of 0.75. Objectives with a higher aperture can be used in conjunction with the above universal condenser only in the equipment version with an integrated aperture iris stop and/or the screw-on darkfield attachment 1.2–1.4 Oil (424218-0000-
000).
•
•
(3) Setting transmitted-light darkfield
•
Set KÖHLER illumination by analogy with transmitted-light brightfield. However, use the objective with the highest aperture instead of the 10x objective.
Turn the turret disk of the universal condenser to position D and swivel in the condenser front lens.
Remove the eyepiece from the tube (or replace it with the auxiliary microscope) and check the centering of the darkfield stop in the exit pupil of the objective. If the central darkfield stop D in the universal condenser is outside of or decentered to the exit pupil of the objective, and if the exit pupil is not homogeneously dark, the darkfield stop must be recentered.
•
After centering, remove both SW 1.5 screwdrivers from the condenser.
Leaving the two Allen screwdrivers unintentionally in the motorized universal condenser does not pose a risk to the drives of the condenser as these instantly switch off when driving against a mechanical resistance. After having removed the source of the resistance, you can continue using the condenser as usual.
Since the apertures of objectives with an integrated aperture iris stop are too high for transmitted-light dark field, the aperture iris stop must be closed to the limit aperture of 0.65.
The darkfield method has been perfectly set, when the background of the field of view appears as dark as possible.
114 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
•
•
•
Reinsert the eyepiece in the tube.
If the height of the dark-field condenser is set correctly and sensitively, it is possible to reduce any brightening in the field of view left, and the luminous-field diaphragm image appears almost perfectly in focus.
Finally, match the size of the luminous-field diaphragm to the size of the field of view.
Fig. 4-56 Centering the darkfield stop on the universal condenser
Dark field microscopy requires specimens to be considerably cleaner than in other techniques.
Fingerprints, dirt or dust particles, in particular, have negative effects, since they brighten the background of the field of view and decrease the contrast of the object image.
B 46-0046 e 12/05 115
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
(4) Use of the achromatic-aplanatic universal condenser 0.9 H/0.8-0.9 DF
For brightfield applications, refer to Section 4.9.1.
The requirements for the use of the universal condenser dependent on the objective used are given in the table below.
2.5x … 5x
10x … 40x
40 x … 100x
Without brightfield front lens H
With brightfield front lens H
With brightfield front lens H
Without darkfield front lens D, with wide-field DF slider for 2.5x-5x
With darkfield front lens D
With darkfield front lens D, with darkfield attachment 1.2-1.4 Oil,
Objective with adjustable iris stop
•
•
•
To swivel the brightfield front lens H into the
optical path, set lever (4-57/4) in position I
(darkfield front lens D swiveled out). To swivel it out, set lever in position 0.
Before swiveling the darkfield front lens D in or out, it is necessary to pull out the wide-field DF
slider (4-57/5) from the universal condenser
(outer click-stop position). To swivel this lens in,
set lever (4-57/8) in position 0. To swivel it out, additionally push locking button (4-57/7) and
set the lever in position I.
For immersion applications, screw the darkfield attachment 1.2-1.4 Oil onto the darkfield front lens and apply Immersol 518 F
®
immersion oil.
Fig. 4-57 Achromatic-aplanatic universal condenser 0.9 H/0.8-0.9 DF
116 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.3 Setting transmitted-light phase contrast
The phase contrast technique is ideal for examining thin, unstained specimens, e.g. culture cells. The human eye is unable to see phase differences (differences in refractive index and thickness) between the different cell components.
The phase contrast technique uses the optical modulators "phase stop and phase ring" as well as the interference procedures during formation of the intermediate image in order to change the small phase differences into intensity and color differences that are visible to the eye.
The high-intensity, direct light components are attenuated with the optically defined ring channel "phase stop and phase ring" and given a constant phase shift. The indirect light components diffracted at different cell components, however, bypass this optical channel and are influenced in phase by the refractive index and the thickness differences in the specimen.
In the intermediate image plane, the thus differently influenced partial beams interfere with each other and are amplified or attenuated - depending on the phase position. This interference results in image contents displaying differences in intensity and color that can be perceived by the human eye.
− Phase-contrast objectives with phase rings Ph 1, Ph 2 or Ph 3 for different average numerical apertures that can also be used in brightfield without any restriction.
− Universal condenser with turret disk containing centerable phase stops Ph 1, Ph 2 and Ph 3 for different average numerical apertures.
− The phase stop on the universal condenser swiveled into the light path must agree with the corresponding label on the objective, e.g. Ph 1.
(3) Setting transmitted-light phase contrast
•
•
Swivel phase-contrast objective, e.g. the one labeled with Ph 1, into the light path.
On the turret disk of the universal condenser, swivel in the phase stop labeled like the phase-contrast objective, e.g. Ph 1.
•
To check centering and congruence of the bright phase stop (in the condenser) with the dark phase ring (in the objective), remove one eyepiece from the tube and replace it with the auxiliary microscope.
Use the correction device on the auxiliary microscope to focus on the phase stop and the phase ring in the exit pupil of the objective.
To check the centration, you can also use the Bertrand lens slider PH. However, this is possible only if there is no camera path deflection installed on the left side of the microscope stand.
B 46-0046 e 12/05 117
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Fig. 4-58 Centering the phase stop on the universal condenser
•
to recenter the bright phase stop until complete congruence with the dark phase ring is
•
Finally, remove the auxiliary microscope from the tube and reinsert the eyepiece.
Normally, however, centering is not required, since the phase stops are factory-centered and centering is retained even if the universal condenser is removed from the condenser carrier and attached again.
To enhance the image contrast, an interference wide-band filter, green 32 x 4, can be inserted in the color glass holder (requires color glass holder for 32 mm filter size).
Perfect phase contrast is only achieved if the bright phase stop (in the condenser) and the dark phase ring (in the objective) are exactly congruent in the
illumination beam path (4-59/B).
Fig. 4-59 Centering the phase stop (bright ring in condenser) to the phase ring
(dark ring in objective)
118 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.4 Setting transmitted-light differential interference contrast (DIC)
The transmitted-light DIC method is a contrast method alternative to polarization allowing contrasty, 3D presentation of transparent specimen details.
The light linearly polarized by a polarizer is split into two partial beams by a birefringent prism. The beams pass two closely adjacent places of the specimen and thus are subject to different path differences caused by differences in refractive index and thickness of the specimen. Afterwards, both partial beams are recombined in a second birefringent prism and, after having passed the analyzer, they have the same vibration direction. Thus, both partial beams can interfere with each other in the intermediate image with the different path differences being converted to different gray values (intensities). A compensator
λ (fullwave plate) then converts the gray values to colors.
− Objectives offered together with the DIC equipment, e.g. Plan-Neofluar objectives
− Objective nosepiece with slot for DIC slider
− DIC slider appropriate for the used objectives
− Condenser with turret disk containing DIC prisms (DIC I, DIC II, DIC III)
− Analyzer module D DIC P&C (424921-0000-000) in reflector turret. Only this module may be used on the Axio Imager as it provides a homogeneous field of view in combination with the special condenser modules I, II and III.
− Preferably a rotary mechanical stage
(3) Setting transmitted-light DIC
•
slider snaps in reliably.
•
•
•
On the reflector turret (4-60/3), swivel in the analyzer module D DIC (4-60/8).
Swivel in the suitable DIC prism I, II or III (digit label on turret disk of condenser (4-60/6)).
Adjust luminous-field diaphragm and aperture diaphragm (4-60/5) in accordance with the rules of
KÖHLER illumination.
•
•
Turn knurled screw (4-60/2) on DIC slider (4-60/1) to adjust optimum contrast. When adjusting the
DIC slider symmetrically around its mid-position, the specimen details can be imaged in such a way that they appear three-dimensionally as if they were raised or dented.
If desired, you can push the compensator
λ (4-60/7) into the slot for the analyzer slider above the
nosepiece to obtain colored DIC contrast.
For 63x objectives, in addition to the DIC sliders HR (High Resolution), another type, HC (High Contrast), is available, which is to be used depending on the type of specimen and examination to be performed.
The DIC method employs polarized light and therefore is disturbed by birefringent objects, such as films, in the light path between polarizer and analyzer, as are occasionally used in conjunction with histological sections. The same applies to acrylic glass culture chambers, if the chamber bottom is made of plastic. In these cases, it is advisable to use chambers with bottom plates of glass to avoid loss of optical performance.
B 46-0046 e 12/05 119
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
1 DIC slider
2 Knurled screw
3 Reflector turret
4 Objective on nosepiece
5 Sliding button for aperture diaphragm
6 Condenser with DIC prism and polarizer
7 Compensator
λ
8 Analyzer module
Fig. 4-60 Components required for transmitted-light DIC
120 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.5 Setting transmitted-light polarization for orthoscopic observation
Magnified viewing, e.g. of a thin section, in polarized light is called orthoscopy (Greek: orthos = straight; skopein = seeing), because illumination is by "straight" light rays, which – with the aperture diaphragm largely closed – are parallel to the microscope axis.
4.9.5.1 Detecting birefringence
(1) Use
The technique of transmitted-light polarization is used for specimens that change the state of polarization of light. These specimens, such as crystals, minerals or polymers, are called birefringent. When these birefringent substances are viewed between crossed polarizers (polarizer
⊥ Analyzer), they appear brightened, while their surroundings remains dark.
Birefringent substances are identified by the fact that they show four bright and four dark positions while being rotated through 360° between crossed polarizers. Dependent on birefringence, thickness as well as orientation of the specimen, interference colors ranging from gray (mostly with biological specimens) to white, yellow, red and blue appear in this process. These interference colors can be of the first or any higher order.
− Phototube Pol
− Achromatic-aplanatic universal condenser Pol
− Strain-free objectives
− Polarizer D (rotatable or fixed) (4-61/3)
− Analyzer slider (4-61/1) or analyzer module D
(4-61/4) in reflector turret (only on microscopes
equipped with a reflector turret)
•
•
•
Set the microscope as for transmitted-light bright field according to KÖHLER (see Section
Center rotary stage Pol (4-61/2) and objectives
(if not done yet – see Section 3.26.3).
Swivel polarizer (4-61/3) into the light path and,
if it is rotatable, turn it to 0°.
Fig. 4-61 Components for transmitted-light polarization
B 46-0046 e 12/05 121
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
•
the reflector turret. The field of view appears dark due to the polarizers being crossed.
If you work with the analyzer slider on the Axio Imager, swivel the reflector turret to a blank turret position.
•
Move the specimen feature you want to examine into the field of view and rotate it with the rotary stage Pol through 360°. As a rule, while being rotated between crossed polarizers, birefringent
(anisotropic) specimens will now show the variations in color and intensity described above. However, optically anisotropic substances may also remain dark, if an isotropic direction, e.g. of optically uniaxial or biaxial crystals, is oriented parallel to the direction of observation.
4.9.5.2 Determining the vibration direction n
γ
'
(1) Use
The determination of the vibration directions of n
γ and nγ' (vibration direction with the absolutely or relatively highest refractive index) and n
α and nα' (vibration direction with the absolutely or relatively lowest refractive index) in relation to the morphological directions, e.g. of crystal surfaces, crystal needles or fibers, provides an important criterion for recognition. It is also used for the diagnosis of biocrystals
(e.g. gout, pseudo-gout).
Fig. 4-61 Determining the vibration direction n
γ
'
by the example of a synthetic fiber
− Phototube Pol
− Achromatic-aplanatic universal condenser Pol
− Strain-free objectives
− Rotary stage Pol
− Polarizer D (rotatable or fixed)
− Analyzer slider or analyzer module D in reflector turret (only on microscopes equipped with a reflector turret)
122 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
The microscope has been prepared as decribed in Section 4.9.5.1 (3).
•
Rotate the rotary stage Pol with the specimen, e.g. a synthetic fiber, until the specimen appears as dark as possible. In this position, the fiber extends parallel to one of the two directions of the crosslines reticle.
•
Next, turn on the rotary stage Pol by 45°, until the longitudinal axis of the fiber is oriented NORTH-
EAST (NE) – SOUTH-WEST (SW) (Fig. 4-61). In this position, the specimen shows the maximum
brightness (diagonal position) and may appear in any color.
Push in the full-wave compensator
λ.
•
Like the specimen, the compensator
λ is a birefringent object, though one with a defined path difference of 550 nm and the principal vibration direction n
γ definitely oriented in NE-SW direction
By moving compensator
λ into the light path, the specimen changes its color. The kind of color change depends on the orientation of the specimen (NE-SW or NW-SE).
The changes in color are based on optical interference. The interference colors (path differences) in both diagonal positions (NE-SW and NW-SE) of the specimen must be compared in this connection.
The path difference results from the superposition (interference) of the vibration direction of the specimen with the vibration direction of the compensator
λ.
The greater path difference occurs, if the vibration direction of the specimen with the absolutely or relatively highest refractive index (n
γ or nγ') is parallel to the principal vibration direction of compensator
λ. The specimen will then appear, for instance, greenish-blue (4-61/2
)
.
The smallest path difference occurs, if the vibration direction of the specimen with the absolutely or relatively lowest refractive index (n
α or nα') is perpendicular to the vibration direction of the compensator
λ. The specimen will then appear, for instance, in yellow (4-61/3
)
.
B 46-0046 e 12/05 123
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Fig. 4-62 Schematic diagram of the color charts according to Michel-Lévy
(4) Conclusions
The grayish-white color appearing first in the
bright position in the above example (4-61/1
) corresponds to a path difference of 150 nm according to the Michel-Lévy color chart (Fig.
When the compensator
λ is brought into the light path, the non-birefringent "surroundings" of the synthetic fiber appear in a dark red color, which corresponds to the path difference of the compensator of 550 nm (1 st
order interference color for the path difference of 550 corresponds to 1
λ).
If the vibration direction (n
γ or nγ') of the birefringent specimen to be examined is parallel to the principal vibration direction (n
γ) of the compensator
λ, i.e. in NE-SW direction, the path difference of the specimen (e.g. grayish-white:
150 nm) and the path difference of the compensator
λ (red: 550 nm) add up. This results in a color change of the specimen from grayish white to greenish-blue (resulting path difference = 700 nm).
If the vibration direction of the specimen to be examined is perpendicular to the principal vibration direction of the compensator
λ, i.e. in NW-SE direction, the path difference of the specimen (e.g. grayishwhite: 150 nm) is subtracted from the path difference of the compensator
λ (red: 550 nm). In this case, the interference color of the specimen visibly changes from grayish-white to orange (resulting path difference = 400 nm).
Color charts according to Michel-Lévy are available under Cat. No. 42-312.
124 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
For the accurate measurement of path differences, measuring compensators are required. These compensators reset or compensate the path difference produced by the specimen to zero (1 st
order black).
While for the methods described above, the addition position or additionally also the subtraction position is of interest, for measurements exclusively the subtraction position is relevant.
Path differences in the specimen can assume very small values (1/50
λ or 10 nm) and very great values
(more than 10
λ or approx. 5500 nm and more) and with that determine the compensator appropriate for the measurement.
To find the appropriate compensator follow this procedure:
•
•
•
•
•
Set the microscope as for transmitted-light polarization (refer to Section 4.9.5.1 (3)).
Position the specimen feature to be examined directly under the center of the reticle.
Restrict the size of the aperture to a value of about 0.2.
Rotate the rotary stage Pol until the specimen is in extinction position, i.e. it appears entirely dark. In this position, activate the 45
° click stop.
Rotate the stage once (by 45
°), so that the specimen is in diagonal position (lightened up).
The intensity of the interference or color produced by the specimen allows the following conclusion to be drawn:
− If more or less strong interference colors appear on the object, the path difference ranges approximately between 1/2
λ and about 5 λ.
The suitable compensator is:
Tilting compensator B 0-5
λ (Accessory 000000-1115-700).
− If the object-side color changes from light gray/white to a strong interference color, when a compensator
λ (473704-0000-000) is inserted in the compensator slot, the path difference is (1/4 ...
1/2)
λ.
The occurrence of the color change effect possibly requires the assessment in two specimen positions lying 90
° apart. For this purpose, rotate the centered stage (by 2 click stops).
The suitable compensator is:
Tilting compensator B 0-5
λ (Accessory 000000-1115-700) or the compensation method according to DE SENARMONT up to 1
λ using the Senarmont compensator 546/4 nm (Accessory 000000-
1115-695).
The compensation method according to DE SENARMONT requires the use of the rotatable analyzer (000000-1076-680).
B 46-0046 e 12/05 125
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
− After insertion of the compensator λ and rotation of the specimen by 90° (2 click stops), the interference color remains to be white, in this case, however, it is a "higher-order white" and thus the path difference is > 5
λ.
The compensator appropriate for this application:
Tilting compensator K 0-30
λ (Accessory 000000-1115-698)
− A dark gray as appearing interference color suggests very small path differences (λ/10 or 54.6 nm).
The compensator appropriate for this application:
Rotary compensator Brace–Köhler
λ/10 (Accessory 000000-1115-703).
•
Push the compensator into the slot as far as it will go.
For measurement preparation and measurement procedure, use the enclosed operating instructions.
126 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.5.4 Transmitted-light circular polarization contrast
(1) Use
Unlike standard polarization contrast, circular polarization contrast does not show any dark (extinction) positions that depend on the angle of rotation (azimuth) of the specimen relative to polarizer or analyzer.
This means, that the image will always look the same while you rotate the stage, as there are no bright and dark positions. With optical anisotropy, all transparent specimens show the interference colors that are characteristic to them.
− Phototube Pol
− Achromatic-aplanatic universal condenser Pol
− Strain-free objectives
− Rotary stage Pol
− Polarizer D (rotatable or fixed)
− Circular polarization equipment D, ACR, with λ/4 plate, rotatable for transmitted light, including reflector module
λ/4, ACR, P&C (427703-9901-000)
− Analyzer slider
• Center rotary stage Pol and objectives (if not done yet - see Sections 3.26.3, 3.26.4).
• For the further settings, for the time being do not use a specimen.
• Push the analyzer slider (4-63/6) into the light path.
the reflector turret.
at full light intensity, assess the extinction (darkening) of the field of view without specimen.
If the extinction is not optimal, correct the position of the vibration direction by slightly and sensibly
turning on the polarizer mount (for this, use the adjusting slots (4-63/4) on the bottom of the
polarizer), until the extinction is maximum. Normally, however, this will not be necessary, as the position of the vibration direction of the polarizer is factory-adjusted.
• Then, swivel the top part of the circular polarizer D (4-63/2) into the light path.
(dark-gray field of view) (lever points 45° to the right).
B 46-0046 e 12/05 127
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Fig. 4-63 Components for circular polarization contrast
• Do not view an (anisotropic) specimen unless the above adjustment has been done.
• Place the specimen to be examined onto the stage.
The specimens appear constantly and independently of the stage rotation in their specific interference color, which depends on the material, specimen thickness and orientation.
To obtain a contrasty image at high objective magnifications (from approx. 20x), reduce the illumination aperture to a value of between 0.15 and 0.20, i.e. close the aperture diaphragm accordingly.
The effect of the
128 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.6 Setting transmitted-light polarization for conoscopic observation – determining the optical character of crystals
For the classification (and thus identification) of crystalline matter, the examination of the interference image in the objective pupil delivers more valuable information than that obtained by viewing the specimen itself. The interference image becomes visible in the eyepiece, if an additional optical system
(fixed or focusing Bertrand lens or, on the basic version, the auxiliary microscope or diopter) is used.
In contrast to orthoscopy, this technique is called conoscopy, because here ideally the specimen is illuminated through a widely open cone. In practical microscopic work, this means that the condenser front lens (0.9 or 1.4) must be in the light path, the aperture diaphragm fully open, and the objective, too, should be a high-aperture type.
(1) Use
The determination of the optical character of transparent and weakly absorbing crystals serves to diagnose crystals. This method is also termed conoscopy. Its main application is the classical mineral microscopy. It allows, however, also synthetic crystals, industrial minerals and plastics (e.g. films) to be identified and characterized.
− Stand with installed phototube Pol or with Bertrand lens slider or with the tube lens turret with integrated Bertrand lens optics
− Strain-free objectives
− Achromatic-aplanatic universal condenser 0.9 H
− Rotary stage Pol
− Polarizer D (rotatable or fixed)
− Analyzer slider or analyzer module D in reflector turret
The phototube Pol can be installed on all Axio Imager stand types.
B 46-0046 e 12/05 129
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
(3) Setting the microscope for conoscopy with the phototube Pol
In the case of uniaxial crystals, the most favorable orientation for conoscopic viewing is obtained with those specimen features (e.g. of a thin section) that in orthoscopic viewing change the brightness as little as possible. In this case, the direction of viewing and the optical axis are
±parallel. The same refers also to biaxial crystals, if they are viewed along or approximately in the direction of one of the two optical axes.
• Set the microscope as for transmitted-light
polarization (see Section 4.9.5.1 (3)).
•
Put the specimen onto the stage and focus on it.
•
Switch phototube Pol to visual observation, if necessary. To this end, pull out the push-pull
rod on the left side (4-64/3).
•
On phototube Pol, push in the front push-pull
rod (4-64/2) on the right side to move the
reticle into the light path.
Fig. 4-64 Axio Imager mit montiertem
Fototubus Pol
•
Move a selected crystal to the center of the reticle.
•
Swivel in the 40x, 50x or 100x objective and, if necessary, refocus the specimen using the focusing drive.
•
Check the centering of the objective by rotating the microscope stage. Recenter it, if necessary.
•
Turn the front push-pull rod (4-64/2) to close the luminous-field stop so far that only the selected
specimen feature remains visible. This is to prevent that the axial figure of the crystal to be examined is not overlapped by axial figures of adjacent crystals. This way, it is possible to mask specimen features of up to 10 µm diameter.
•
Move the Bertrand lens on the phototube Pol into the light path. To this end, push in the rear push-
the axial figure, turn this push-pull rod.
(4) Setting the microscope with Bertrand lens slider or tube lens turret with Bertrand lens optics for conoscopy of large-size specimens
• Set the microscope as for transmitted-light polarization (see Section 4.9.5.1 (3)).
•
•
Put the specimen onto the stage and focus on it.
Swivel in the 40x, 50x or 100x objective and, if necessary, refocus the specimen using the focusing drive.
•
Check the centering of the objective by rotating the microscope stage. Recenter it, if necessary.
•
Close the luminous-field diaphragm so far that only the selected specimen feature remains visible.
130 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
•
Push the Bertrand lens slider into the light path. Focus the axial figure by means of the lever of the slider, or, on the tube lens turret, swivel the position with the Bertrand lens into the light path, and focus the axial image by turning the focusing wheel.
(5) Evaluation
Crystalline anisotropic specimens can be classified in optically uniaxial and biaxial specimens, each with
"optically positive" or "optically negative" character.
Uniaxial crystals show a black cross, if the optical axis is oriented parallel to the viewing direction.
Dependent on the degree of birefringence and the specimen thickness concentrically arranged
colored interference fringes (the so-called isochromats) might appear (see Fig. 4-65, second row).
This cross remains closed, when you rotate the stage. It can be located inside or outside the imaged objective pupil, depending on the position of the section.
With optically biaxial crystals, the cross resolves into two dark hyperbola branches (the so-called
isogyres) depending on stage rotation, which are surrounded by colored interference patterns depending on the amount of birefringence and specimen thickness (suggestive of the figure "8").
Inserting a compensator
λ (473704-0000-000) or λ/4 (473714-0000-000) or a wedge compensator 0-4 λ
(000000-1140-663) in the compensator slot with the initial state of the axial figure being as illustrated in
figure, thus allowing differentiation in "optically positive" and "optically negative".
λ plate
(white
→ blue
→ yellow)
Optically uniaxial Optically biaxial
Positive Negative Positive Negative
+ = blue
– = yellow
Quartz wedge
(Direction of motion at insertion)
λ/4 plate
(position of black spots)
Fig. 4-65 Determining the optical character
Ò Direction of
Ó movement
B 46-0046 e 12/05 131
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
If the positions of the section are more unfavorable, i.e. the center of the cross of optically uniaxial or the isogyre of optically biaxial specimens is outside the objective pupil, the optical character can be assessed as follows:
•
The specimen is optically uniaxial if the isogyres are straight and run (related to the reticle) parallel through the pupil.
•
The specimen is optically biaxial if the black isogyres are curved lines traveling across the pupil on a circular path.
Axial figures can often be better represented by circular polarization. Especially the determination of the axial angle of optically biaxial specimens (more or less the space between isogyres) succeeds with a much greater unambiguity. The optical character can also be determined by inserting the compensator
λ (6 x 20) into the compensator slot.
For all reflected-light contrasting techniques, the compensators 6x20 must be removed from the light path (slot for compensators).
(1) Application
The reflected-light brightfield microscopy is the simplest and most widely used microscopy technique which is used to examine opaque samples or specimens e.g. polished sections or wafers.
For a true-to-object imaging, indirect ray bundles, i.e. ray bundles diffracted and scattered on the specimen details, are of major importance in addition to the so-called direct ray bundles. The higher this portion of indirect rays (aperture), the more realistic the microscope image will be, according to ABBE's rule.
The incoming, bundled light from the reflected-light illuminator is reflected by a neutral-colored beam splitter. Then it passes to the objective which focuses the beams onto the specimen surface (so-called condenser function). The objective collects the light reflected by the object and generates the intermediate image of the microscope together with the tube lens, which is then observed visually or can be documented objectively.
− Axio Imager MAT with connected and adjusted HAL 100 halogen illuminator.
− H P&C reflector module in reflector turret, 6x20 compensator mount with darkfield stop for reflected light (424706-0000-000) or 4-position modulator turret.
132 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
(3) Setting reflected-light brightfield according to KÖHLER
− The microscope has been started properly as described in Section 3.
− The microscope has been switched on.
•
Switch on the halogen lamp for reflected light using the reflected-light/transmitted-light toggle switch
(4-1/36) on the microscope stand.
Depending on the existing equipment, the microscope contains a 6x20 compensator mount or a 4position modulator turret for setting the contrasting techniques. The 6x20 compensator mount can be used for both brightfield and darkfield. For C-DIC and TIC examinations, additionally the corresponding
6x20 slider is required. Refer also to Section 4.9.10.
•
•
•
•
•
The 4-position modulator turret has a combined brightfield/darkfield position (H/D) as well as three additional positions for C-DIC (C1, C2) and TIC (TIC).
•
When using the 6x20 compensator mount, remove the 6x20 slider, if necessary. When using the 4position modulator turret, set the H/D position.
Swivel the reflector turret into brightfield position H.
Adjust light-intensity control (4-66/5) on microscope stand.
Place a contrasty reflected-light specimen on the stage.
Turn nosepiece (4-66/7) to swing in 10x objective (yellow ring, see also Section 2.5).
•
Use focusing drive (4-66/6) to focus on the specimen. In doing so, always focus away from the
specimen if possible to avoid any collision between objective and specimen.
Remove the reflected-light diffusion disk. Turn adjusting screws of HAL 100 halogen illuminator to focus and center the image of the lamp filament in the exit pupil of the objective. For that, either pull out the adjusting aid or remove one eyepiece from the binocular tube. Afterwards, push the adjusting aid in again or reinsert the eyepiece. Move the reflected-light diffusion disk into the light path again.
•
wheel.
•
•
Turn the focusing drive (4-66/6) to refocus on the edge of the luminous-field diaphragm (4-66/B) and
•
•
Then, open the luminous-field diaphragm (4-66/4) so that it just disappears from the field of view
To set the aperture diaphragm (image contrast), remove one eyepiece from the binocular tube and look into the tube with your naked eye or insert the auxiliary microscope in place of the eyepiece.
•
Center the aperture diaphragm with the centering screws (4-66/1) and, for specimens with average
contrast, adjust the size of the aperture diaphragm to about 2/3 to 4/5 of the exit pupil diameter of
the objective (4-66/E) by means of knurled wheel (4-66/2).
In most applications, this aperture diaphragm setting provides optimum contrast at almost ideal resolution, and is therefore the best compromise for the human eye.
•
Finally, reinsert the eyepiece, refocus with the coaxial coarse and fine focusing drive (4-66/6) and
adapt the image brightness to the specimen to be examined.
Never use the aperture diaphragm to control the image brightness, but use the light intensity
control (4-66/5), or swivel attenuation filters of the 2-position filter wheels into the light path!
B 46-0046 e 12/05 133
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Fig. 4-66 Microscope settings on Axio Imager MAT in reflected-light brightfield
134 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
(1) Application
The incident light darkfield technique is used to examine specimens that do not only have reflective surfaces of different reflectivity (ideal brightfield objects), but feature scratches, cracks, pores, or in a nutshell, deviations in plane surfaces. All these light-scattering details shine brightly in the darkfield whilst the reflective plane surfaces stay dark.
− Axio Imager MAT with connected and adjusted HAL 100 halogen illuminator.
− Epiplan-Neofluar, EC Epiplan-Neofluar, Epiplan objectives additionally labeled with "HD".
− Reflector module DF P&C, 6x20 compensator mount with darkfield stop for reflected light (424706-
0000-000) or 4-position modulator turret
reflected-light
•
•
•
•
•
•
diaphragm should just disappear from the field of view to avoid reflections.
Rotate the reflector turret to move the DF P&C reflector module into the light path.
When using the 6x20 compensator mount, remove the 6x20 slider, if necessary. When using the
4-position modulator turret, set position H/D.
Rotate the nosepiece to swing in the darkfield objective (HD).
Fully open the aperture diaphragm and remove any neutral-density filter from the light path.
Place the specimen on the stage and focus on it.
B 46-0046 e 12/05 135
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
4.9.9 Setting reflected-light DIC and reflected-light C-DIC
(1) Application
The reflected-light DIC and reflected-light C-DIC technique (DIC = Differential Interference Contrast,
C-DIC = Differential Interference Contrast in Circularly polarized light) serves for producing high-contrast images of phase specimens, i.e. those specimens which only change the phase of the light in contrast to amplitude specimens.
− Axio Imager MAT with connected and adjusted HAL 100 halogen illuminator.
− Rotary mechanical stage 75x50/240°
− EC Epiplan-Neofluar, Epiplan objectives additionally labeled with "DIC" or "Pol".
− DIC slider, that specially fits the objective that is engraved with magnification and aperture on the top of the slider or 6x20 C-DIC slider (in connection with the reflector module C DIC P&C), 6x20 compensator mount or 4-position modulator turret).
•
•
•
•
(3) Reflected-light DIC, preferably for Epiplan 5x - 100x and LD-Epiplan 10x - 100x objectives
•
diaphragm should just disappear from the field of view to avoid reflections.
Rotate the reflector turret to move the C DIC P&C reflector module into the light path.
Rotate the nosepiece to swing in the objective position with DIC slider slot.
Push the DIC slider into the slot on the nosepiece (above the objective).
•
Place the specimen on the stage, focus on it and rotate the mechanical stage until the specimen structure of interest appears at maximum contrast.
To optimize the contrast, you can turn the knurled screw on the DIC slider.
•
•
•
Prepare the microscope for reflected-light brightfield.
Switch the C DIC P&C reflector module into the light path.
Push the C-DIC slider 6x20 (4-67/3) into the
6x20 compensator mount (4-67/4) or rotate the
4-position modulator turret (4-68/6) at turret wheel (4-68/5) to swing in the desired C-DIC
position (C1 or C2).
C1 for 5x … 20x objectives;
C2 for 50x … 100x objectives.
When using objectives of 50x or higher, it is advisable to use the reflected-light diffusion disk.
Fig. 4-67 6x20 compensator mount with
6x20 C-DIC slider
136 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
•
Place the specimen on the stage, focus on it
and rotate the control wheel (4-68/1) on the
•
(4-68/1) until the specimen detail of interest is
visible at maximum contrast. Rotating the stage is no longer necessary. If both line marks are
facing each other (4-68/3), this corresponds to
the mid-position. Rotation by 45° in both directions is possible).
The contrast can be optimized by turning
corresponds approximately to the extinction position (best contrast).
Fig. 4-68 4-position modulator turret
B 46-0046 e 12/05 137
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
4.9.10 Setting reflected-light TIC
(1) Application
The reflected-light TIC technique (microinterferometry; TIC = Total Interference Contrast in circularly polarized light) can be used to image and measure object structures available in different azimuths.
− Axio Imager MAT with connected and adjusted
HAL 100 halogen illuminator.
− EC Epiplan-Neofluar, Epiplan objectives additionally labeled with "DIC" or "Pol".
− 6x20 compensator mount or 4-position modulator turret
− 6x20 TIC slider with accompanying C DIC P&C reflector module.
Fig. 4-69 6x20 TIC slider
•
•
Place the specimen (e.g. a step-shaped object) on the stage and prepare the microscope as
described in Section 4.9.7 for reflected-light
brightfield.
Rotate reflector turret to swing C DIC P&C reflector module into the light path.
•
Push 6x20 TIC slider into 6x20 compensator mount (4-69/4) or rotate turret wheel (4-68/5) of
fringes appear. Turn setscrew (4-69/2) of the TIC slider or the modulator turret to shift the black
interference fringe until it appears to be in the center of the field of view.
•
The interference fringes can be shifted by means of setscrew (4-69/2) of the TIC slider or the
modulator turret.
The step height is determined subsequently according to the following formula: d
= n
∆
2
=
λ b
2 a where: d = step height in nm n = refractive index of the environment, usually air (n = 1)
∆ = path difference a = spacing of interference fringes b = offset of interference fringes at the step
λ = wavelength of the illumination in nm
138 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
The values for a and b (see Fig. 4-70) are
determined using the eyepiece reticle or the micrometer eyepiece.
If you are working with white light (without interference filter),
λ = 550 nm must be used.
When using interference filters, their center wavelength applies.
The measured path difference is aperturedependent and decreases with increasing illumination aperture.
b a
Fig. 4-70 Interference fringe pattern
Accordingly, the following correction values must be taken into account depending on the objective used:
5x/0.15 1.0057
10x/0.25 1.0161
10x/0.30 1.0236
20x/0.4 1.0436
20x/0.50 and 50x/0.75 1.0718
50x/0.60 1.1111
50x/0.75 and 100x/0.75 1.2038
50x/0.80 1.2500
50x/0.90 and 100x/0.90 1.3929
10x/0.95 1.5241
Table 1: Aperture-dependent correction
Example:
a = 11 mm
λ = 550 nm b = 5 mm
Objective 20x/0.50 d
=
λ ⋅ b
⋅ k
2 a
=
550 nm
⋅
5 mm
22 mm
⋅
1 .
0718
=
134 nm
B 46-0046 e 12/05 139
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Caution:
•
If the step and the environment are of different materials, the phase jumps inherent to the material must be taken into account. Whilst the phase jump for all non-conductors is 180° and for semiconductors only deviates slightly from 180°, i.e. the measuring error is negligible, the measured values may be falsified for metals on glass for example. The phase jumps in table 2 calculated for vertically incident light and compact material are to serve as recommended values because it can be assumed that the phase jumps depend on the layer thickness and the angle of incidence of the light.
Accurate determination of the thickness is only possible by coating the entire object with a homogeneous layer and then measuring the path difference.
•
If the layers or steps are transparent such as silicon dioxide on silicon, the interference fringes may change their color and the determination of the interference order then become problematical. This can also be remedied by additionally coating the surface with a homogenous layer.
Material
Phase jump
φ
Copper 140.0° determination of the thickness:
Gold 142.5°
∆
−
2
δφ
2
Bismuth 151.0°
Φ =
140
°
,
Φ glass
=
180
° therefore part of the phase jump
Zinc 159.0°
Platinum 160.0°
δφ
2
=
20
°
or
λ
18
=
30 nm
Aluminum 160.0° without taking the phase jump inherent in the material great.
Chromium 165.0°
Carbon 160.0°
Graphite 165.0°
Silicon 177.0°
Glass 180.0°
Table 2: Calculated phase jumps for compact material and vertically incident light
140 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.11 Setting epi-fluorescence
CAUTION
To reduce the transmission, use an FL attenuator, discrete (423616-0000-000 or 423617-
0000-000). The gray filters mounted in the 2-position filter wheels (428300-0000-000 or
428301-0000-000) are not permanently stable.
The epi-fluorescence technique enables high-contrast images of fluorescent substances to be displayed in typical fluorescence colors. In the epi-fluorescence microscope, light generated by a high-performance illuminator reaches the exciter filter (band pass) through a heat-absorbing filter. The filtered, short-wave excitation light is reflected by a dichroic beam splitter and focused on the specimen via the objective. The specimen absorbs the short-wave light and then emits the long-wave fluorescence light (Stoke’s law), which is now gathered by the objective and transmitted by the dichroic beam splitter. Finally, the rays pass a barrier filter (long pass/band pass), which only allows the long-wave light from the specimen to be transmitted.
Exciter and barrier filters must be perfectly matched. They are arranged in a reflector module FL P&C together with the corresponding dichroic beam splitter.
− Recommended objectives: Plan-Neofluar or Fluar objectives (UV excitation)
− Reflector module FL P&C in reflector turret
− Mercury vapor short-arc lamp HBO 100 for reflected-light illumination
− Halogen illuminator HAL 100 for transmitted-light illumination
Before using the epi-fluorescence technique, make sure to align the mercury vapor short-arc lamp by means of the adjusting aid as described in Section 3.23.3. Re-alignment may be necessary depending on the operating time.
(3) Setting
The first epi-fluorescence setting is considerably simplified if you begin with the Plan-Neofluar objective
20x/0.50 and a strongly fluorescing specimen. You may also use demonstration specimens first.
Before setting epi-fluorescence, make sure to remove compensator
above the nosepiece, which may have been left there from a previously performed transmittedlight DIC examination.
•
•
•
•
Switch on halogen illuminator HAL100.
Swivel in Plan-Neofluar objective 20x/0.50.
First, swivel condenser turret to brightfield position H (or phase contrast Ph) and set the specimen feature to be examined.
For the time being, keep the light path in the reflected-light illuminator blocked by reflected-light shutter RL (rear right on microscope stand) (indicator LED is lighting).
B 46-0046 e 12/05 141
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
Fig. 4-71 Components for epi-fluorescence on Axio Imager
•
Switch on the HBO 100 mercury vapor short-arc
lamp (4-71/1) and allow it to warm up to
operating temperature for about 15 minutes.
•
On the reflector turret (4-71/2), select the
reflector module FL P&C containing the desired fluorescence filter combination (depending on the desired kind of excitation) and swivel it in.
•
Remove the reflected-light shutter RL from the light path in the reflected-light illuminator and close the transmitted-light shutter TL.
•
Remove one eyepiece from the tube and set the aperture diaphragm with your eye. For that,
open the aperture diaphragm (4-71/4) until it
clears the full exit pupil of the objective. Center the aperture diaphragm to the exit pupil using
the two centering screws (4-71/3 and 5), if
necessary.
•
Reinsert the eyepiece in the tube and close the
luminous-field diaphragm (4-71/7) until it is
visible in the field of view.
•
Use the two centering screws (4-71/6 and 8) to
center the luminous-field diaphragm to the edge of the field of view.
•
Open the luminous-field diaphragm until it just disappears from the field of view or, if there is a risk of specimen bleaching, close it so far that it is visible in the field of view.
• Finally, refocus on the specimen and optimize described in Section 3.23.3. Adjust the collector in such a manner that the field of view is illuminated as evenly as possible when using the short-wave excitation reflector module. When you use long-wave excitation modules, correction of the collector position is not required.
142 B 46-0046 e 12/05
OPERATION
Axio Imager Illumination and contrast methods Carl Zeiss
4.9.12 Setting reflected-light polarization – Detection of bireflection and reflection pleochroism
(1) Use
Incident-light polarization presents another contrasting option for polished sections of ore minerals, coals, ceramic products, certain metals and metal alloys, as these specimens often show a varied reflection behavior in linearly polarized light depending on the orientation of the crystals and specimen features.
The illumination light is linearly polarized by the polarizer and directed through the objective onto the specimen surface, where it is reflected. Here, the light rays experience structure-dependent path differences or polarization-optical rotations, which on passing the analyzer appear as different gray-scale values. A compensator with lambda plate enables the conversion from gray to color contrast. Important!
However, you must not use a 6x20 compensator for this purpose!
With objectives of very low magnification, a rotatable
λ/4 plate arranged in front of the objective
(Antiflex cap) permits the reflections to be eliminated even with "dark" specimen surfaces, which otherwise would be unavoidable.
− Stand with installed and aligned HAL 100 halogen illuminator.
− Epiplan-Neofluar Pol, EC Epiplan-Neofluar Pol, Epiplan Pol objectives.
− Reflector module DIC P&C or DIC Rot I P&C in reflector turret; or reflector module Pol P&C plus analyzer slider; or analyzer slider plus polarizer slider.
reflected-light
•
•
•
•
Set the microscope for reflected-light brightfield as described in Section 4.9.7.
If you use the objective position with DIC slider slot, remove the possibly inserted DIC slider.
also swing in reflector module Pol P&C and insert the analyzer slider in the corresponding slot. When
into the corresponding slots. If you use the fixed versions of these sliders, the polarizer is oriented in
EAST-WEST direction and the analyzer in NORTH-SOUTH direction.
Put specimen onto stage, set the desired magnification, focus and observe the specimen in the now adjusted polarization contrast while rotating the stage.
If the specimen features show variations in brightness and color, when the stage is rotated, the specimen is said to have bireflection.
If the specimen has a weak bireflection only, it is advisable to use the analyzer with rotatable lambda plate.
Pleochroism can be made out by color variations occurring in the specimen while rotating the stage (with the reflected-light polarizer moved into the light path and the analyzer moved out).
B 46-0046 e 12/05 143
OPERATION
Carl Zeiss Illumination and contrast methods Axio Imager
If the microscope is equipped with the phototube Pol, the following settings are required for this contrast method:
– The Bertrand lens must be inactive (rear push-pull rod (4-72/6) on right side pulled out).
– The luminous-field diaphragm must be open (front push-pull rod (4-72/5) turned
counterclockwise as far as it will go).
– The crossline reticle must be inactive (front push-pull rod (4-72/5) pulled out).
Fig. 4-72 Components for reflected-light polarization
144 B 46-0046 e 12/05
Axio Imager
5
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Instrument care
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Carl Zeiss
Care of the Axio Imager is restricted to the following operations:
•
•
Eliminate contaminations on the instrument according to the rules for prevention of accidents.
•
•
Switch off the instrument each time after use and place the instrument cover on it to protect it from dust and humidity.
Do not set up the instrument in a humid room; maximum humidity < 75%.
•
Remove dust and loose dirt on visible optical surfaces with a brush, blower brush, cotton bud, optics cleaning tissue or cotton cloth.
Remove water-soluble dirt (coffee, cola, etc.) by blowing on it and wiping it off with a lint-free cotton cloth or a cloth moistened with water to which you may also add a mild detergent.
•
Wipe off stubborn, oily or fatty dirt, e.g. fingerprints and immersion oil, with a cotton swab or lint-free cotton cloth and the optics cleaning solution L.
The cleaning solution consists of 90 vol% gasoline and 10 vol% isopropanol (IPA). The individual components are also known as:
Gasoline: Medical alcohol, petrolether
Isopropanol: 2-Propanol,
Dimethylcarbinol,
2-Hydroxypropane
Clean optical surfaces by polishing in circles, starting in the middle and moving to the edges (only use slight pressure).
When using the microscope in humid climatic zones, proceed as follows:
•
Store the instrument in bright, dry and well-ventilated rooms with a humidity of < 75 %. Optical components and accessories that are particularly susceptible to fungus growth, e.g. objectives and eyepieces, should be stored in a dry closet.
The risk of fungus growth on opto-mechanical instruments invariably exists in the following conditions:
− Relative humidity > 75% and temperatures between +15 °C and +35 °C for more than three days.
− Installation in dark rooms without air ventilation.
− Dust deposits and fingerprints on optical surfaces.
B 46-0046 e 12/05 145
Carl Zeiss
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Instrument maintenance Axio Imager
•
•
•
•
Makes sure the line voltage available agrees with the operating voltages required.
Check the power cable and plug for defects.
If any damage is visible, switch off the instrument. Have the instrument repaired only by a qualified technician.
Check the reading of the operating hour meter of the power supply to ensure that the maximum operation time of the mercury vapor short-arc lamp is not exceeded.
Always pull the power plug before changing the fuses.
Fig. 5-1 Changing fuses
• compartment is on the back of the microscope and contains two T 5A / 250V fuses.
•
•
•
•
Pull the power plug.
Pull out fuse holder (5-1/2) frontward. Use a
small screwdriver, if necessary.
Remove the fuses from the holder and insert new fuses.
Push the fuse holder back into the fuse
compartment (5-1/1) as far as it will go.
Connect the power plug.
The motorized Axio Imager model is powered through the power supply 230. The fuse compartment of the power supply is on the rear panel containing two T 6.3A / 250V fuses.
Change the fuses by analogy with the manual stand.
146 B 46-0046 e 12/05
Axio Imager
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Troubleshooting Carl Zeiss
5.3 Troubleshooting
Problem Cause
Shadows or inhomogeneous image brightness in the field of view; the field is not entirely visible.
The vis/doc push-pull rod on the phototube is not in the correct position (intermediate position).
No DIC effect
The nosepiece with objective has not clicked into place.
The condenser has not been set correctly.
The aperture diaphragm has not been set correctly.
Remedy
Move the vis/doc push-pull rod to the correct position (end position).
Turn on the nosepiece with the objective until it clicks into place.
Set the condenser correctly
(adjustment, centering);
Set the aperture diaphragm correctly (centering, aperture);
The luminous-field diaphragm has not been set correctly.
The filter has not been inserted correctly in the filter mount.
The polarizing elements are not in the light path.
Set the luminous field-diaphragm correctly (centering, opening);
Insert the filter correctly in the filter mount.
Move the polarizing elements into the light path and set them to crossed position.
DIC slider in the light path. DIC slider in the light path. Double images in bright-field examinations
Low resolving power and poor image contrast.
The aperture diaphragm has not been opened to the correct size.
Set the aperture diaphragm as per the 2/3 rule or the specimen
The condenser has not been focused correctly and front lens
0.9 not swiveled in / out correctly.
Focus the condenser and swivel front lens 0.9 correctly in or out;
Wrong cover slip thickness for transmitted-light objectives corrected for 0.17 mm cover slip thickness.
Specimen slide placed upside down.
Use of no or non-specified immersion oil with immersion objectives.
Use standardized 0.17 mm cover slips.
Turn over the specimen slide; the specimen side should be on top.
Use immersion oil 518 F
®
from
Air bubbles in the immersion oil. Remove the bubbles by applying new oil.
B 46-0046 e 12/05 147
Carl Zeiss
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Troubleshooting Axio Imager
Problem Cause
Low resolving power and poor image contrast.
Immersion oil on the front lens of a dry objective.
The cover-slip correction ring has not been set to the correct coverslip thickness.
Remedy
Clean the front lens of the dry
Set the correction ring to the correct thickness; see p. 23.
Dirt or dust on the optical surfaces of objectives, eyepieces, condensers or filters.
Clean the respective optical
Asymmetrically blurred images, e.g. one side in focus, one side not in focus.
The condenser has not been set correctly.
Set the condenser correctly;
Greater focus differences after objective change.
Left and right fields of view cannot be combined to one image.
The nosepiece has not clicked into place.
Turn on nosepiece until it clicks into place (click-stop).
The specimen has not been clamped in position on the stage.
Correctly insert the specimen in the specimen holder and clamp it.
The focusing eyepieces have not been set correctly.
The objective has not been screwed in completely.
The tube lens has either not been inserted or inserted superfluously.
The interpupillary distance of the binocular tube has not been set correctly.
Set the focusing eyepieces to the appropriate ametropia; see p. 32.
Screw objective fully in as far as it will go.
Insert the tube lens or remove the superfluous tube lens.
Set the interpupillary distance correctly; see p. 33.
Eye-fatiguing microscopy.
The focusing eyepieces have not been set correctly.
The interpupillary distance of the binocular tube has not been set correctly.
The focusing eyepieces have not been set correctly.
The image brightness is not acceptable.
Binocular tube optically / mechanically out of alignment.
Set the focusing eyepieces to the appropriate ametropia; see p. 32.
Set the interpupillary distance correctly; see p. 33.
Set the focusing eyepieces to the appropriate ametropia; see p. 32.
Adjust the lamp voltage or insert the conversion filter.
Have it checked / repaired by
Microscopy Service.
148 B 46-0046 e 12/05
Axio Imager
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Troubleshooting Carl Zeiss
Problem Cause
Dirt or dust in the field of view. The condenser has not been focused correctly and front lens
0.9 not swiveled in/out correctly.
The aperture diaphragm opening is too small.
Remedy
Focus the condenser and swivel front lens 0.9 correctly in or out;
Set the aperture diaphragm as per the 2/3 rule or the specimen
Dirt or dust on the optical surfaces of the objectives, eyepieces, condensers, filters or specimens.
Clean the optical surfaces of the
affected components; see p. 145.
The 12V 100W halogen lamp does not function although the on / off switch is set to "on".
The power plug has not been connected to the power outlet.
Connect the power plug to the power outlet considering the power requirements of the instrument.
12V 100W halogen lamp has not been installed.
12V 100W halogen lamp is defective.
Failure to use the specified 12V
100W halogen lamp.
The fuses are defective.
Insert 12V 100W halogen lamp; see p. 50.
Replace 12V 100W halogen lamp; see p. 50.
Use the specified 12V, 100W
Replace defective fuses;
The electronic module is possibly defective.
Have electronic module checked by service and replaced, if
Use another power outlet. The power outlet does not supply line voltage.
The 12V 100W halogen lamp is flickering, the light intensity is unstable.
End of average service life of
12V 100W halogen lamp.
Incorrectly installed or broken power cable.
Replace 12V, 100W halogen lamp; see p. 50.
Connect the power cable correctly or replace it.
The pins of the 12V 100W halogen lamp haven not been inserted correctly in the receptacle.
Insert the pins of the 12V 100W halogen lamp correctly in the receptacle; see p. 50.
B 46-0046 e 12/05 149
Carl Zeiss
5.4
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Spares, consumables and tools
Spares, consumables and tools
Axio Imager
12V 100W halogen lamp
HBO 103 W/2 mercury vapor short-arc lamp
SW 1.5 ball-headed screwdriver *
SW 3 ball-headed screwdriver *
SW 4 offset screwdriver
Eyepiece eyecup
*
Dust cover for nosepiece
Dust cover for eyepiece sockets
Immersion medium Immersol 518 F
®
;
Oiler, 20 ml
Bottle, 100 ml
Bottle, 250 ml
Cleaning tissue, 300 sheets
G-fuse inserts (5 x 20 mm);
T 5 A/H / 250V 2x
(Stand, manual)
G-fuse inserts (5 x 20 mm);
T 6.3 A/H / 250V 2x
(Power supply 230 for motorized stand)
Dust cover set M
Dust cover set L
Set of cover caps
(included in 434303-0000-000 and
434304-0000-000)
380079-9540-000 For HAL 100 illuminator
380301-9350-000 For HBO 100 illuminator
000000-0460-470 For changing condensers and stages
000000-0069-551 For changing tubes and illuminators
000000-0015-278 For removing the carrying handle
444801-0000-000 Recommended for low-light techniques to suppress reflections
462981-0000-000
000000-0168-373
444960-0000-000
444962-0000-000
444963-0000-000
462975-0000-000 For cleaning optical surfaces
434303-0000-000
434304-0000-000
434302-0000-000
For closing instrument openings not in use
For oil immersion applications n
D
= 1,518
Protects the integrated power supply from excessive load
Protects the power supply from excessive load
For covering the instrument when it is not in use.
For covering eyepieces
* Included in the tool kit that is supplied with the instrument in tool bag (451892-0000-000).
150 B 46-0046 e 12/05
Axio Imager
CARE, MAINTENANCE, TROUBLESHOOTING AND SERVICE
Requesting service Carl Zeiss
All repairs of mechanical, optical or electronic components inside the instrument and of the electrical components of the Axio Imager microscopes may only be performed by Carl Zeiss service staff or specially
authorized personnel.
To ensure optimum setting and trouble-free function of your microscope over a longer period of time, we recommend that you enter into a service/maintenance agreement with Carl Zeiss.
For subsequent orders or when service is required, please get in touch with your local Carl Zeiss representative.
For additional information, contact us at [email protected] or visit us on the Internet at http://www.zeiss.de
B 46-0046 e 12/05 151
APPENDIX
Carl Zeiss List of abbreviations Axio Imager
6 APPENDIX
6.1 List of abbreviations
AC Alternating
ACR
Br.
Automatic Component Recognition
Suitable for eyeglass wearers cod. Coded
CSA Canadian Association
D
D / DF
Cover slip thickness
Darkfield d Diameter (e.g. of filters)
DIC Differential Contrast
DIN Deutsches Institut für Normung (German Standards Institute) doc Documentation
EC European
EN European
EMC
FL Fluorescence foc. focusing
H Brightfield
HAL Halogen
HBO
ICS
Mercury vapor short-arc lamp for fluorescence
Infinity Color-Corrected System
IEC International
IP International Protection (protection by instrument casing)
ISO International
LED man.
Light Emitting Diode manual
MC Microscope mot. n
D motorized
Refractive index for D line (sodium)
Ph Phase
PL Plan
Pol Polarization
P&C Push&Click
R Right (drive knob to the right of the mechanical stage)
RL Reflected
152 B 46-0046 e 12/05
APPENDIX
Axio Imager List of abbreviations Carl Zeiss
SW
T
Wrench size across flats
Slow-blow (fuse type)
TV Television
UL Underwriter
UV ultraviolet
VDE Verband Deutscher Elektrotechniker (Association of German Electricians) vis visual
V obj
Magnification of objective
W 0.8"
W-PL
Whitworth-type thread 0.8"
Xenon short-arc lamp XBO
B 46-0046 e 12/05 153
APPENDIX
Carl Zeiss Index Axio Imager
6.2 Index
Page
Numbers
2TV tube mot ........................................................................................................................................75
3200 K ..................................................................................................................................................66
A
Adjusting aid ...................................................................................................................................64, 65
Ambient conditions ...............................................................................................................................26
Ametropia .............................................................................................................................................32
Analyzer ......................................................................................................................................121, 127
Analyzer module .................................................................................................................................122
Analyzer slider .........................................................................................................................64, 65, 122
Aperture diaphragm ..............................................................................................................68, 112, 142
B
Beam splitter .........................................................................................................................................42
Binocular tube ...........................................................................................................................30, 65, 72
Birefringence .......................................................................................................................................121
Brightfield .....................................................................................................................68, 111, 117, 132
C
Camera path deflection, left ............................................................................................................64, 70
Camera path deflection, right ................................................................................................................65
CAN/USB converter ...............................................................................................................................79
Care ....................................................................................................................................................145
Checks, performing .............................................................................................................................146
Coarse focusing control .............................................................................................................64, 67, 68
Color filter carrier ..................................................................................................................................62
Color glass filter mount .......................................................................................................................118
Color temperature, 3200K...............................................................................................................64, 66
Compensator mount .............................................................................................................................37
Condenser.......................................................................................................29, 35, 64, 68, 70, 73, 111
Condenser carrier ..............................................................................................................35, 64, 67, 113
Connecting to power ............................................................................................................................47
Connections ..........................................................................................................................................55
Conoscopy ..........................................................................................................................................129
Consumables.......................................................................................................................................150
Contrast Manager .................................................................................................................................86
Coupler plate ........................................................................................................................................31
Crystals, determination of the optical character ...................................................................................129
D
Darkfield .............................................................................................................................................114
DIC......................................................................................................................................................136
DIC Differential Inference Contrast ......................................................................................................136
DIC prisms .............................................................................................................................................36
Differential interference contrast..........................................................................................................119
Diffusing glass, reflected-light..........................................................................................................64, 69
154 B 46-0046 e 12/05
APPENDIX
Axio Imager
Diffusing glass, transmitted-light......................................................................................................64, 66
Dimensions............................................................................................................................................26
E
Ergonomic drive.....................................................................................................................................57
Eyecups .................................................................................................................................................33
Eyepiece ................................................................................................................................................65
Eyepiece reticle ......................................................................................................................................32
Eyepieces.......................................................................................................................24, 29, 31, 32, 64
F
Filter set.................................................................................................................................................40
Filter wheel, 2-position ....................................................................................................................38, 39
Filter wheel, 2-position, discrete.....................................................................................64, 65, 66, 70, 72
Fine focusing control .................................................................................................................64, 67, 68
FL attenuator, discrete .........................................................................................................64, 66, 70, 73
Fluorescence........................................................................................................................................141
Focusing drive........................................................................................................64, 67, 68, 70, 73, 113
Front lens ................................................................................................................................68, 73, 111
Fuses, changing ...................................................................................................................................146
H
Halogen illuminator ...............................................................................................................................48
Halogen illuminator HAL 100.................................................................................................................64
Halogen lamp ........................................................................................................................................50
HBO 100 illuminator ........................................................................................................................53, 54
I
Illumination and contrast methods.......................................................................................................111
Illuminator HBO 100 ..............................................................................................................................64
Installation.............................................................................................................................................29
Instrument care ...................................................................................................................................145
Instrument description ...........................................................................................................................12
Instrument maintenance ......................................................................................................................146
Instrument safety.....................................................................................................................................6
Intended use..........................................................................................................................................12
Intermediate tube ..................................................................................................................................45
Interpupillary distance............................................................................................................................33
K
KÖHLER.......................................................................................................................................111, 119
L
LED illuminator ......................................................................................................................................51
Light intensity ..................................................................................................................................64, 66
Light Manager .......................................................................................................................................80
Light sources .........................................................................................................................................27
Line voltage .....................................................................................................................................26, 27
LM Set...................................................................................................................................................69
Luminous-field diaphragm .........................................................................................64, 67, 70, 111, 142
B 46-0046 e 12/05 155
APPENDIX
Carl Zeiss Index Axio Imager
M
Maintenance .......................................................................................................................................146
Mechanical stage.................................................................................................................29, 56, 59, 64
Mechanical Stage 75x50 mot. CAN .......................................................................................................77
Menu structure......................................................................................................................................87
Mercury vapor short-arc lamp ..............................................................................................................142
Microinterferometry ............................................................................................................................138
Modulator turret, 4-position ........................................................................................................136, 138
Multidiscussion equipment ....................................................................................................................45
O
Objective nosepiece .................................................................................27, 28, 35, 64, 68, 70, 111, 133
Objectives......................................................................................................................23, 29, 35, 57, 60
ON/OFF switch.................................................................................................................................64, 68
Operation..............................................................................................................................................26
Operation and function controls ......................................................................................................63, 70
Orthoscopy..........................................................................................................................................121
Overall view.....................................................................................................................................10, 11
P
P&C reflector module ............................................................................................................................40
Phase contrast .....................................................................................................................................117
Phototube .............................................................................................................................................34
Phototube 30°/25 mot with 2 camera ports ...........................................................................................75
Polarization .........................................................................................................................121, 129, 143
Polarizer ................................................................................................................................62, 121, 122
Polarizer for transmitted light ..........................................................................................................64, 67
R
Reflected light .....................................................................................................................132, 141, 143
Reflected-light brightfield ....................................................................................................................132
Reflected-light C-DIC ...........................................................................................................................136
Reflected-light DIC ..............................................................................................................................136
Reflected-light illuminator....................................................................................................................142
Reflected-light polarization ..................................................................................................................143
Reflected-light shutter .........................................................................................................64, 66, 68, 69
Reflected-light TIC ...............................................................................................................................138
Reflector module FL ...............................................................................................................................40
Reflector turret ........................................................................................................27, 28, 37, 64, 68, 70
Requesting service ...............................................................................................................................151
Reticles..................................................................................................................................................24
Rotary stage Pol.....................................................................................................................................59
S
Safety......................................................................................................................................................6
Smoothness...........................................................................................................................................58
Spares .................................................................................................................................................150
Specimen holder..............................................................................................................................29, 59
Stage carrier ..................................................................................................................35, 56, 57, 59, 60
Stage focusing.................................................................................................................................27, 28
Stage lift, quick-action .....................................................................................................................70, 74
156 B 46-0046 e 12/05
APPENDIX
Axio Imager
Stage lowering, quick-action............................................................................................................70, 73
Stage micrometers.................................................................................................................................24
Stop slider .................................................................................................................................64, 65, 70
Switching off .........................................................................................................................................74
Switching on .........................................................................................................................................74
System overview ....................................................................................................................................15
T
Technical data .......................................................................................................................................26
TFT display.................................................................................................................................43, 70, 85
Toggle switch, transmitted/reflected light illuminator .............................................................................69
Tools ...................................................................................................................................................150
Torque...................................................................................................................................................58
Touchscreen ....................................................................................................................................70, 85
Transmitted light .........................................................................................111, 114, 117, 119, 121, 129
Transmitted/reflected light toggle switch ...............................................................................................64
Transmitted-light shutter .....................................................................................................64, 66, 68, 69
Troubleshooting ..................................................................................................................................147
Tube..............................................................................................................................30, 31, 45, 64, 70
Tube lens turret .....................................................................................................................................31
U
Universal condenser.........................................................................................................36, 68, 114, 117
Unpacking .............................................................................................................................................29
V
Vibration direction ...............................................................................................................................122
Viewing height ......................................................................................................................................33
W
Warranty .................................................................................................................................................9
Weight ..................................................................................................................................................26
B 46-0046 e 12/05 157
APPENDIX
Carl Zeiss Industrial property rights Axio Imager
6.3 Industrial property rights
Instruments, instrument components or methods described in this manual are protected by patents and registered utility models:
US6276804
US6392796
US5015082
CH691699
GB2306585
US6123459
DE29821694
158 B 46-0046 e 12/05
Download
Public link updated
The public link to your chat has been updated. You may view and manage your chat links after registration here.
Document public link
Advertisement
Key features
Transmitted-light brightfield, darkfield, phase contrast, and DIC microscopy
TFT display
Various button configurations
Biological and materials microscopy
Frequently asked questions
To set up transmitted-light brightfield microscopy, follow the steps outlined in Section 4.9.1 (3) of the user manual. Ensure the microscope is switched on, and the toggle switch for halogen illuminators is set to transmitted light. Then, use the vertical control of the condenser drive to lower the condenser until the edge of the luminous-field diaphragm appears in focus. Center the luminous-field diaphragm image, and open the diaphragm until its edge just disappears from the field of view. Finally, adjust the aperture diaphragm to about 2/3 to 4/5 of the diameter of the exit pupils of the objectives.
To configure the button functions, you'll need to access the User page on the navigation button bar. From there, you can choose between Standard Settings and Personal Settings. To edit the button assignments, you'll need to enter an administrator password.
The difference between a biological and a materials microscope lies in their intended use and the types of specimens they are designed to handle. A biological microscope typically focuses on examining biological samples, such as cells, tissues, and organisms. It features features like a wide range of objectives and illumination methods suited for biological observation. A materials microscope, on the other hand, is designed for examining materials such as metals, ceramics, and polymers. It often has specialized objectives and illumination methods designed for analyzing the structure and properties of materials.
