Practical Parasitology

Microscopy


Kohler Illumination

Principle

The main objective of using Kohler Illumination is to acheive the optimal resolving power
of the microscope. It is achieved when the illumination system is aligned with the optics
and focused on the specimen.

Purpose

The purpose is to control the light path from lamp to objective. The result will be timely
and accurate identifications with the least amount of eye strain.

Method


  1. Focus on the specimen.
  2. Close the field diaphram fully.
  3. Focus the field diaphram by moving the condenser up or down until the closed field
    diaphram is sharply in focus.
  4. Center the diaphram with the centering screws on the condenser.
  5. Open the field diaphram until the outer edge just disappears out of view.
  6. Adjust the condenser diaphram by removing and ocular. While looking down the empty
    tube, adjust the condenser diaphram to close the light opening by 1/3.

Calibration of Ocular Micrometer

Principle

Laboratory diagnosis of intestinal parasites in most cases is dependent on the size of
the organisms. It is therefore, a requirement that laboratories undertaking the recovery
and identification of intestinal parasites have available a calibated ocular micrometer

Purpose

Each technologist must comply with CAP, LPTP, and Laboratory Licensing regulations regarding
the calibration of ocular micrometers. The differences between each microscope and each
technologists interpupillary distances make it necessary to calibrate each microscope
under all objectives they may work with.

Method

  1. With the ocular micrometer in place, focus on the calibrated stage micrometer starting
    with the low power objective.

  2. Adjust the stage micrometer so that the "0" line of the ocular micrometer is exactly
    superimposed over the "0" line of the stage micrometer. See Diagram below.

  3. Look as far as possible along both sides until tou see two lines exactly over one
    another. At higher magnifications it may be necessary due to the thickness of the lines
    to line up the marks to the left or right of the marks.
  4. Count the number of ocular divisions between the "0" and the point where the second set
    of lines is superimposed. On the stage micrometer count the number of 0.1mm divisions
    between the "0" and the second set of superimposed lines.
  5. The calculation is as follows

1 ocular unit = stage units/ocular units X 1000
In our case example 1 ocular unit = 0.40/70 X 1000 = 5.7 microns

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