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Prior to Köhler illumination critical illumination was the predominant technique for sample illumination. Critical illumination has the major limitation that the image of the light source (typically a light bulb) falls in the same plane as the image of the specimen, i.e., the bulb filament is visible in the final image.
August Karl Johann Valentin Köhler (4 March 1866 – 12 March 1948) was a German professor and early staff member of Carl Zeiss AG in Jena, Germany.He is best known for his development of the microscopy technique of Köhler illumination, an important principle in optimizing microscopic resolution power by evenly illuminating the field of view.
The technique most commonly used in microscopy to optimize the light pathway between the condenser (and other illumination components of the microscope) and the objective lens is known as Köhler illumination. The maximum NA is limited by the refractive index of the medium between the lens and the sample.
Critical illumination or Nelsonian illumination is a method of specimen illumination used for transmitted and reflected light (trans- and epi-illuminated) optical microscopy. Critical illumination focuses an image of a light source on to the specimen for bright illumination.
Major techniques for generating increased contrast from the sample include cross-polarized light, dark field, phase contrast and differential interference contrast illumination. A recent technique combines cross-polarized light and specific contrast-enhanced slides for the visualization of nanometric samples. [citation needed]
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The system incorporates oblique angle, pre-aligned Kohler illumination. [4] The resulting high signal-to-noise image enables direct observation of nanoscale sample elements. The dual mode fluorescence module is a transmitted light fluorescent technique that enables real time observation of both fluorescent and non-fluorescent sample elements.