Light Microscope

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LAB: THE LIGHT MICROSCOPE AND CELL STRUCTURE INTRODUCTION The microscope is an instrument designed to observe objects that are too small to be seen with the naked eye. In this class, you will use a compound light microscope which functions by passing light through the object being viewed and focuses this beam of photons into an image using optical lenses. Your microscope has the capability of magnifying an object 1000 times its normal size and its resolving power can distinguish two objects that are separated by as little as 0.5 um. You should be able to identify and describe the function of the various components of a typical compound light microscope as shown below.

4) Always focus an image by moving the lens away from the slide. This way you won’t accidentally grind the objective lens into the slide while looking through the microscope. 5) When changing objective lenses, rotate the nose piece by turning it from its circular base. Never rotate the nose piece by grasping an objective lens as this will loosen the lens. 6) Never use the oil immersion objective lens (white ring at its base, labeled 100X) unless directed to do so by the instructor. It requires that a special oil be placed between the slide and the lens; without it, you may damage the objective lens. 7) When finished viewing, always store the microscope with the scanning objective in place. This minimizes the chance of an objective lens grinding into the stage during transport to/from the cabinet. Learning to focus an image with the microscope may seem a bit tricky at first, but the task will become second nature in a very short time. The diagram below illustrates the distance between slide and objective lens for a properly focused image. WORKING DISTANCE BETWEEN TIP OF OBJECTIVE AND SLIDE WHEN AN OBJECT IS IN FOCUS Scanning Objective 4X Low Power Objective 10 X 40 X 25 mm 8.3 mm microscope slide 0.5 mm High Power Objective

Microscopes are expensive instruments; they can best be maintained by following a few important guidelines: 1) Always carry the microscope to/from the storage cabinet with two hands, one on the arm and one on the base. Set it gently on the counter (pretend it’s a baby’s head) so that you don’t jar the lenses out of place. 2) Never tilt the microscope, the ocular may fall out. 3) To avoid scratching the lenses, never clean them with anything other than lens tissue.

The following procedure should enable you to focus an image with little difficulty. 1) Make sure the ocular and objective lenses are clean; use lens tissue if necessary. 2) Open the diaphragm to allow the maximum amount of light to pass from the light source, through the lenses, to your eye. 3) Make sure the low power objective is in place. (If the object to be viewed is very large, start with the scanning objective.) 4) Make a wet mount (see below) and place the slide on the stage, securing it with the stage clips. Make sure the object to be viewed is in the center of the light beam. 5) As you watch from the side, use the coarse focus knob to move the low power objective all the way down (or until it is almost touching the slide).

6) Now, looking through the objective lens, use the coarse focus knob to raise the objective lens until the object comes into view. 7) Once the object is in focus, use the fine focus knob to sharpen the focus. 8) Adjust the diaphragm to get the best lighting. 9) If further magnification is desired, you will need to use the high power (40x) objective. As you watch from the side, rotate the nose piece so that the high power objective slides into position. The objective lens will be very close to the slide but it should not hit the slide. 10) Now, looking through the objective lens, use the fine focus knob to sharpen the focus. Never use the coarse focus knob when the high power objective is in place because a small turn of the coarse focus knob in the wrong direction will grind the objective into the slide. 11) Adjust the diaphragm to get the...
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