5: MEASURING WITH A MICROSCOPE
- Page ID
- 157064
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Describe the relationship between magnification and field of view in compound light microscopes.
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Identify how different objective lenses (4x, 10x, 40x, 100x) affect the visible area and level of detail.
- Estimate the actual size of specimens in micrometers by comparing them to calibrated measurements.
BACKGROUND
Microscopes make it possible to observe objects that are far too small to be seen with the naked eye, such as bacteria, plant cells, or grains of sand. When viewing these microscopic structures, it is important to determine their actual size. This is accomplished through a process called calibration, which allows accurate measurement through the microscope.
The area visible when looking through a microscope is called the field of view, and its size depends on the objective lens in use. Most microscopes include 4x, 10x, 40x and 100x objectives. The higher the magnification the more detail seen, however, the size of the field of view in decreased. For example, if a specimen fills the entire field at 4x magnification, it must be significantly larger than something that fills the field at 40x. This provides a general idea of an object’s size, but more precise measurement requires an established reference.
To make accurate measurements, a micrometer slide is used. This special slide has a fine ruler etched into the glass with units marked in micrometers (µm). One micrometer equals one-thousandth of a millimeter. For context, a human hair is roughly 70 micrometers wide.
Calibration is performed by placing the micrometer slide on the microscope stage and viewing it under each objective lens. By counting how many micrometer divisions span the field of view at each magnification, it is possible to calculate how much of the specimen is being viewed. This reference makes it possible to estimate the actual size of unknown samples. For instance, if the field of view is known to be 400 micrometers wide and a specimen fills half the field, then its approximate size is 200 micrometers.
Accurate measurement under the microscope plays an important role in many fields. Medical laboratory technicians measure blood cells to help diagnose disease. Environmental scientists study the size of microorganisms in water or soil to help in determining contamination. Forensic scientists compare the length and diameter of hairs or fibers found at crime scenes. In clinical settings, the size of a microorganism or cell is an essential clue for identification. Two species may appear visually similar but differ slightly in length or width. Even a few micrometers of difference can indicate a different organism or health condition.
In this lab, you will observe several microscopic specimens under various magnifications. By drawing each sample at different powers and comparing them to the calibrated micrometer slide, the actual size of each specimen can be estimated in micrometers.
MATERIALS (Per Student)
1 Micrometer slide
1 Prepared letter “e” slide
1 Clean microscope slide
1 Strand of hair
2 Pieces of Tape
1 Prepared WBC slide
1 Prepared RBC slide
METHODS/PROCEDURES
1. Place the micrometer slide on the microscope stage and center it so the etched scale is clearly visible in your field of view.
2. Focus carefully using the lowest objective lens. Once the image is sharp, draw what you see. Make sure your drawing includes the entire circular field of view and shows how much of the micrometer scale is visible. Label the magnification used.
3. Repeat step 2 for each of the remaining objective lenses. You should end up with four micrometer drawings, one for each magnification.
4. Tape a strand of your hair onto a clean microscope slide. Run the single strand of hair down the middle of the microscope slide horizontally then tape the hair to the edges of the slide. Be sure the tape is not in your field of view and lays flat on the slide. You do not need much tape. One 1 X 1cm piece on each edge is sufficient.
5. Observe and draw the prepared sample slides. For each sample, the magnifications to be drawn are listed in the table below. Always start with the 4x objective but only draw what you see the field of view at the magnifications listed in the table.
*It is important that you carefully observe and draw exactly what you see and your
drawing is proportional to the full field of view—show how much of the circle the
sample takes up. Inaccurate drawings will affect your results
6. Once all drawings are complete, use your micrometer drawings to estimate the actual size of each sample by comparing the sample drawing to the micrometer drawing made at the same magnification.
7. Record the estimated size of the samples in the results section.
*Remember that the full micrometer scale represents 1 millimeter (1 mm),
which is equal to 1,000 micrometers (µm).
The following is to be completed during lab then turned in on Canvas as a PDF
*If you are using an iPad or tablet you will need to take screen shots of your competed work, save the screenshots as one PDF then submit them on Canvas by the due date designated on Canvas.
*You can also print out the entire exercise to bring to lab with you. If you choose to complete the lab on paper, take pictures of the completed results and conclusions sections only, save them as one PDF, then submit to Canvas by the due date designated on Canvas.
MEASURING WITH A MICROSCOPE
NAME ______________________
EXPECTATIONS
What do you think will happen to the size of the field of view as you switch from low power to high power on the microscope?
RESULTS
Draw your observations below. Remember to label underneath the circle with the name of the sample and final magnification.
CONCLUSIONS
1. Which specimen was the most difficult to measure? What challenges did you face, and how did you handle them?
2. Which specimen was the easiest to measure accurately? What made it easier to estimate its size?
3. If two different samples both fill the field of view at different magnifications, what might that tell you about their actual size?
4. How might an incorrect size estimate under the microscope lead to a mistake in diagnosing a patient’s condition?