1.2 Microscopes, Handling, Maintenance

2009 Holcim/Switzerland

Microscopes, Functions, Handling, Maintenance

Microscopy Workshop at Kymore Plant

India, 25 29 May 2009

Bruno Misteli

217.05.2009Microscopy Workshop 2009 2009 Holcim/Switzerland

Parts of the MicroscopeTrinocular tube Eyepieces Analyzer for transmitted and reflected light Daylight filterField of view diaphragm for reflected lightAperture diaphragm for reflected lightLambda or gypsum plateRevolving nosepiece ObjectivesRotating stageSwing out condenserAperture diaphragmStage drive or focusing knobPolarizer for transmitted lightVoltage controlField of view diaphragm for transmitted light


The Field of View Diaphragm

The field of view diaphragm limits, as is indicated by its name, the supply of light to the area to be investigated.

The field of view diaphragm should be centred at a low opening and focused by moving it up or down until its boundaries are well defined.

The field of view diaphragm should be opened as little as necessary, but wide enough to make it disappear out of view.

A minimised opening limits the amount of diffuse light in the system and improves the image contrast.


The Aperture Diaphragm

The aperture diaphragm is used to adjust the brightness of the picture.

The change of voltage supply to the light source is in general not recommended to adjust the brightness, as the colours change when using different voltages.

Closing the aperture diaphragm decreases the amount of light and increases the depth of field and thus reveals internal structures, here observed in polarised transmitted light with a -plate.


Aperture Diaphragm Decentration (in reflected light)

Some microscopes are equipped with an aperture diaphragm decentration device. The use of this device is in general limited to reflected light applications. It can be used to amplify topographic effects. This device is very useful to differentiate between hard and soft phases. A similar effect can also be achieved when using microscopes without this device. An obstacle, like a partly, just slightly inserted lambda plate into the light path can have a similar effect. The identification of soft free lime crystals (left side of the centre) is here much easier possible.

Centred diaphragm Same area observed with a decentred diaphragm


Khler Illumination

a) Field diaphragm not focused, not centred

c) Field diaphragm focused and centred, but diameter too small

b) Field diaphragm focused, but not centred d) Field diaphragm diameter = object field diameter Khler illumination


Uniform Illumination by Lamp Adjustments

a) Direct filament image but decentred

b) Direct filament image in the right position

c) Reflected and direct filament image in the right position


Darkfield and Brightfield Illumination

Darkfield illumination

Darkfield illumination is the classical stereo microscope illumination, which in general does not require sample preparation efforts for observations. Only diffuse reflected light reaches the eye. The colours are identical to the ones observed by the naked eye.

Brightfield illumination is the classical reflected light microscope illumination requiring a co-axial light path. Etching effects (here HF-etching) become visible.

Brightfield illumination (same area as left side)


Darkfield Illumination, External Light Source

When the microscope is equipped with a low magnifying objective such as 4x or 2.5x with a long working distance, similar observations to stereo-microscopes are possible. An external light source is required for such observations. Special care is, however, required not to spill the sample onto the optics underneath.


Microscope Handling

1. Start your investigations always with a low magnification objective.

2. Adjust the eyepiece distance until the pictures within both eyes unite to one single picture.

3. Adjust the crosshair in the eyepiece for the right eye until it is in focus.

4. Focus an object on the stage with the right eye by adjusting the stage position. Start with a low position of the stage and move it slowly towards the objective lens until a sharp picture is achieved.

5. Leave the stage in the same position and adjust the left eyepiece until the object is in focus.

6. The object and the crosshair should now be perfectly in focus, if not, start again at point 2.


What To Do in Case of Blurred Pictures? Blurred pictures are often caused by dust or smears on optical parts. Clean eyepieces with an optical cleaning liquid and a lint free tissue. Never use solvents, as the lens fitting material might be attacked. If no

commercial cleaning liquid is at hand, use distilled water with a trace of liquid soap.

Check the light path for any blockages, like inadvertently half inserted -plate, etc.

If the picture is not in focus on one side, make sure that the surface of the object under investigation is perfectly perpendicular to the optical axis of the microscope.

Use an air brush in case of dust on the polished section. Re-impregnate and re-prepare highly porous samples causing excessive

diffuse reflections.

If an objective appears to be smeared, remove the objective from its socket and check it by looking through from the back side. Clean it with an optical cleaning fluid and check its alignment (see next slide).


Alignment of Objectives

Check whether a particle under the cross hair remains in place while the stage is rotated.

In case this particle describes a circle, adjust the objective by means of its two objective adjustment screws in the revolving nose piece.

Stop rotating the table when the particle is in position 2.

Move the particle by means of the objective adjustment screws to point 3.

Move the particle from position 3 to position 1 by means of the x-y object stage or by manual movement.

Check again by rotating the table and repeat the procedure if necessary.

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How To Maintain a Good Optical Quality

Helpful articles, when fighting dust or smears on optical parts


Treat Your Microscope Carefully

Cover up your microscope when not in use, to protect it from dust. Never expose your microscope to acid vapors. When working with

hydrofluoric vapor etches, make sure your sample is clean and sufficiently blown with a hairdryer before you take it to the microscope.

When focusing problems occur at higher magnifications, go back to lower magnifications to find the focus again. You can thus avoid to damage optics or samples by crashing into them.

If necessary adjust the vertical position of all objectives to the same focus plane by inserting special washer plates.

Do not use oil to lubricate stage drive. Your stage might want to move down by its own weight. Use silicon grease if necessary, which does no harm to plastic parts.


Lets Practice!

Microscopes, Functions, Handling, MaintenanceParts of the MicroscopeThe Field of View DiaphragmThe Aperture DiaphragmAperture Diaphragm Decentration (in reflected light)Khler Illumination Uniform Illumination by Lamp AdjustmentsDarkfield and Brightfield IlluminationDarkfield Illumination, External Light SourceMicroscope HandlingWhat To Do in Case of Blurred Pictures?Alignment of ObjectivesHow To Maintain a Good Optical QualityTreat Your Microscope CarefullyLets Practice!

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1.2 Microscopes, Handling, Maintenance