Proper alignment of the illuminating slit is critical in achieving the optimum results with Hoffman Modulation Contrast microscopy. The following problems and solutions are often encountered when imaging specimens using this technique.

Problem: The image is very low in contrast and is difficult to visualize and/or produces flat photomicrographs also lacking is sufficient contrast.

Solution: This problem can arise from incorrect alignment of the illuminating slit, or when the contrast control polarizer is missing or rotated out of position. Use a Bertrand lens or a phase telescope to re-align the slit while observing its image at the rear focal plane of the condenser. Make certain that the contrast control polarizer is aligned with the vibration direction perpendicular to the polarizer in the slit opening. This is accomplished by rotating the polarizer while observing the slit with the phase telescope. When properly positioned, the image of the slit should appear superimposed over the gray area of the modulator.

Problem: Illumination of the specimen is uneven and the image appears to have dark and/or light overtones.

Solution: This problem may arise for several reasons: (i) the wrong slit plate is being used with the objective, or (ii) the image of the illuminating slit is not properly aligned and superimposed on the gray area of the modulator, or (iii) the overall microscope illumination is not properly adjusted.

(i) Rotate the turret condenser until the matching slit plate is in place in the optical axis. Repeat the illuminating slit alignment procedure with a Bertrand lens or phase telescope.

(ii) Use a Bertrand lens or a phase telescope to adjust the image of the illuminating slit plate making sure it is properly superimposed on the gray area of the modulator.

(iii) Adjust the microscope for Köhler illumination using a brightfield condenser with an adjustable aperture diaphragm.

Problem: The illuminating slit can not be imaged on the gray region of the modulator plate.

Solution: In most cases, the wrong illuminating slit plate is being used in the substage condenser. Make sure the slit plate is matched to the objective and realign the microscope with a Bertrand lens or phase telescope.

Problem: When imaging the back focal plane of the objective, the dark and gray zones are not present in the modulator.

Solution: The wrong objective is being used. Carefully remove the objective and hold it up to a bright light source and peer through the rear lens. The modulator plate should be visible, if it is present and properly aligned in the objective. If not, change to an objective that contains a modulator plate.

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BACK HOFFMAN MODULATION CONTRAST

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