Using the Microscope
Finding objects under the microscope
With a slide on the microscope stage, move the low power objective into place. As you look through the ocular (eyepiece) lens, the circular area you see is the field of view. The coarse adjustment knob is for focusing when the low power objective is in place. Using the coarse adjustment knob, raise the stage until the slide is in focus. It is important that you are careful when raising the stage with the coarse adjustment knob. If you raise it too far, the lens and the slide can collide, damaging both. Be sure to look from the side of the stage to be sure that you have plenty of room. Once the slide is in focus, you can use the fine adjustment knob to make the image even clearer.
INSERT DIAGRAM OF MICROSCOPE
If you need to see more detail, use the medium or high power objective lenses. To do this, follow the steps outlined below:
1. Center the image in your field of view and make sure that it is in focus.
2. Next, move the medium power objective into position. When doing this, make sure that there is enough room for the lens to clear the slide. Notice that the specimen seems much larger under medium power than it did under low power. This is because the medium power lens has magnified it further.
3. The microscope is parfocal, which means that an image focused under low power should remain in focus under medium or high power; although some fine focusing may be required. When viewing an object with the medium or high power lens, always use the fine focus knob. The coarse focus knob moves the stage too much and it is likely that you would damage the lens and/or the slide.
To view the object with the high power lens, first make sure that it is centered and in focus. Then move the high power lens into position and adjust the image using the fine focus if necessary.
If you find that you are unable to see anything on the slide after switching to a more powerful lens, try going back to low power and starting again.
When using the high power lens, you may find that you need to decrease the amount of light reaching the ocular lens. If this happens, the image will appear "washed out" and you will be unable to see some fine detail. In some cases, you may see nothing at all. To decrease the amount of light reaching your eye, simple close the diaphragm (or iris) slightly so that less light is able to get through.
Remember that, even though they appear quite thin to the naked eye, the objects you view with the microscope are three dimensional. Consequently, you cannot focus on the entire object at once. To get a better understanding of what the object looks like, you will need to turn the fine focus knob back and forth to bring different layers of the object into focus. This way, you can develop a more accurate perception of what the object is like in three dimensions.
Depth of Field
When observing a specimen, say, a cell, under the microscope, it is important to remember that the object is three dimensional. When you are observing a portion of the specimen, you are seeing only a layer of the specimen in focus. The depth of field is the amount of an image that is in focus at any one time. To get an accurate idea of how the specimen really appears, it is necessary to focus back and forth so that you can see different 'layers' of the specimen in focus at different times. This is especially important if you are observing cells. Because the depth is limited, you may miss certain important cellular features if you fail to focus back and forth 'through' the cell.
Field of View
The field of view is the lighted, circular area you see when you look through the ocular lens. The diameter of the field of view can be calculated rather easily.
1. With the low power objective in place, put a transparent ruler on the stage so that you can see the millimeter divisions.
2. Focus on the divisions.
3. Move the ruler so that one of the millimeter marks is at the edge of the field of view. Now you can count the number of millimeter divisions you can see across the field. Estimate it to the nearest 0.5 mm. This is the diameter of the field of view. Convert millimeters to micrometers (mm). (1 mm = 1000 mm.)
4. The field of view for both medium and high-power magnification can be determined indirectly by calculating the ratio quotient of the high-power objective lens to the low-power objective lens. Calculate the ratio quotient for each lens using the equation:
| Ratio quotient = | magnification of desired lens |
| magnification of low power lens |
5. Calculate the diameter of the field of view for both the medium and high-power objectives (use micrometers).
| Desired lens diameter = | low-power field diameter |
| ratio quotient |
Preparing a wet mount
A wet mount is used for observing specimens that could dry out as you are observing them, especially living cells. The wet mount keeps the specimen moist so that it does not dry out or shrivel.