OPERATIONAxioscope 5/7/Vario Illumination and contrast methods in transmitted light … ZEISS03/2018 430035-7344-001 874.2 Illumination and contrast methods in transmitted light microscopy4.2.1 Setting up transmitted light brightfield microscopy using the KÖHLER method(1) General principle of operationOf all the optical microscopy methods, transmitted light brightfield microscopy is the one most commonlyused. High-contrast or tinted (stained) samples (e.g. a blood smear) can be examined easily and quickly.For an imaging result which is as true to the specimen as possible, we must not only consider the so-called direct bundled beams but the indirect ones as well, i.e. the beams which diffract and scatter on thesample details. According to ABBE, the image is truer to the specimen when the fraction of the cone oflight is as large as possible.The best performance of the microscope, and especially its objective, is achieved when the condenser,field diaphragm and aperture diaphragm are adjusted in accordance with the KÖHLER illuminationprinciples. These fundamental basic rules for adjusting a microscope are explained in detail insection 6 0 8 H 4.2.1 (3) "Setting up transmitted light brightfield microscopy using the KÖHLER method".(2) Instrumentation for transmitted light brightfield microscopyEvery microscope (except the one with the Vario stand column) is configured to work with thetransmitted light brightfield method.All available condensers (except special condensers like darkfield condensers) can be used for thetransmitted light brightfield method.(3) Setting up transmitted light brightfield microscopy using the KÖHLER method− The Axioscope has been put into operation appropriately (section 6 0 9 H 3).− The Axioscope is turned on.• Adjust the image brightness using the Intensity/LM knob (610HFig. 4-3/1) on the microscope stand.• Put a high-contrast sample into the specimen holder of the mechanical stage.• If condensers with a swiveling front lens are used, swivel these into the beam path with ≥ 10xobjectives and turn the knurled knob for vertical adjustment of the condenser (611HFig. 4-3/3and 6 1 2 H Fig. 4-4/2) to the top stop. Make sure the stop is adjusted to prevent the condenser from liftingout the sample (adjusting the condenser stop, section 6 1 3 H 4.2.1 (4)).• On condensers with revolver/modulator disks: turn the knurled ring (614HFig. 4-4/3) to position H (or B =brightfield).• Bring the 10x objective into position on the nosepiece (615HFig. 4-3/6) and focus the sample using thefocus drive (616HFig. 4-3/2).• Close the field diaphragm (617HFig. 4-3/5) enough to make it visible in field of view (even if blurred)(618HFig. 4-3/A).• Lower the condenser using the knurled knob for vertical adjustment until the edge of the fielddiaphragm appears sharp (619HFig. 4-3/B).
