microscope | Biology Laboratory Manual I (2023)

Things you should be able to explain to someone else after this lab:

• Mixture
• full zoom
• Binocular
• Field of view
• depth of field
• investment phenomenon

The slideshow

Bottom

Many organisms (bacteria) and parts of organisms (cells) that biologists study are too small to be seen by the human eye. We are usingmicroscopesto enlarge samples for our research. The word microscope means "to see small", and the first primitive microscope was made in 1595.

There are several types of microscopes, but mainly you will use:complex optical microscope. This type of microscope uses visible light focused by two lenses, an eyepiece and an objective lens, to view a small sample. Only cells that are thin enough to allow light to pass through will be visible under a light microscope in a two-dimensional image.

The next microscope you will use in the lab isstereoscopicby oneautopsy microscope. This type of microscope uses visible light to view thicker and larger specimens, such as insects, in 3D. Because you are looking at larger samples, the magnification range of a dissecting microscope is less than that of a compound light microscope.

Your instructor will review the parts and functions of the complex light microscopes we will be using throughout the term. Complete the table on the next page to help you remember this important information. You will probably come back to this page often. here it islight microscope imageso you can mark up and take notes.

Rules and instructions for using complex light microscopes

The instructor will discuss the use of the microscope with the class. The relevant steps to follow to apply the right approach will be reviewed. You must follow the step-by-step sequence indicated by the instructor. Even if you are familiar with this type of microscope, you are expected to go through focus review with the rest of the class.

1. Important rules:
   1. Always carry the microscope with both hands: one hand on the microscope arm and the other under the microscope base.
   2. Do not touch the objective lenses (i.e., the ends of the lenses).
   3. When adding or removing slides, keep the lenses in the scanning position and leave the stage.
   4. Always look at the microscope from the side when making large stage height changes.
   5. Lenses should only be cleaned with special lens paper and lens cleaning fluid.
   6. DON'T be a tinkerer: do not disassemble any microscope. Report faults to your instructor.
2. To remove the microscope from the lab cabinet:
   1. First, clean the table area under the microscope; avoid a crowded workplace.
   2. The microscopes are numbered depending on where you sit. Find the number under the bench and use the appropriate microscope.
   3. Carry the microscope with TWO hands.
   4. Secure the power cord, don't let it hang off the table!
3. Putting the microscope away in the laboratory cabinet:
   1. Lower the stage.
   2. Rotate the scan target to the correct place on the stage.
   3. Remove the slide from the scene.
   4. Clean the glass and lens with the special lens cleaner and supplied paper.
   5. Center the stage so it doesn't stick out too much to either side.
   6. Secure the cable by wrapping it under the microscope stage.
   7. Replace the dust cover.
   8. Carry the microscope with TWO hands.
   9. Put the microscope back in the same place you bought it, and be sure to place it within arm's length.
4. To focus the microscope:
   Take a look at the microscope from the side:
   1. Lower the stage as much as possible.
      1. Rotate the targets so that the scan target is facing down the stage.
      2. Adjust the scene so that the aperture (the hole in the center of the scene) is centered.
      3. Place the slide on the stage using the stage clamp to secure it. The stage clamp moves only in the horizontal plane of the stage and secures the slide by touching the lower right corner of the slide.
      4. Center the sample on the slide below the objective by moving the stage.
      5. Move the stage up as far as possible, taking care that the slider does not touch the lens; look from the side to see how high you need to raise the stage.
   2. Look through the glasses:
      1. Use the coarse adjustment knob to move the stage down until the sample is in focus, then sharpen the focus.
      2. Center the sample in the microscope field by moving the stage.
   3. Take a look at the microscope from the side:
      1. Slowly rotate the low magnification (10X) objective to the correct position.
   4. Look through the glasses:
      1. Adjust the sharpness with the coarse adjustment knob if necessary. Generally only minimal adjustment is necessary.
      2. If necessary, center the sample in the field.
   5. Take a look at the microscope from the side:
      1. Rotate the high power target into place.very carefully!
   6. Look through the glasses:
      1. For sharper focus, use the fine adjustment dial only. Re-center if necessary. (If the sample is "disappeared", immediately return to low power and re-center the sample.)
      2. When removing a slide, always set the lenses to the scanning position and lower the stage before lifting the slide.

Part 1: Full extension

increaseis the ratio of the size of the image obtained under the microscope to the actual size of the object. When you say magnification is 10, the image you see under the microscope is ten times larger than what you see with the naked eye. Remember that with a compound light microscope, you magnify with two lenses, so to calculate the total magnification, multiply the objective magnification by the eyepiece magnification. Look under the microscope and use the table below to calculate the total magnification of each lens:

Part 2: The Reversal Phenomenon

Get the "e" slide available in class. Observe the slide with your eyes and then place it under the microscope. Use focus sequence to view the slide at low magnification.

Procedure

1. Draw the letter "e" as it appears when you look at the slide without a microscope.
2. Draw the letter "e" as it appears when you look at the slide under the microscope.
3. Move the slider to the right with the mechanical lever of the stage while looking through the glasses. In which direction does the "e" move?
4. Move the slider away from you using the mechanical lever of the stage while looking through the glasses. In which direction does the "e" move?

Explain the phenomenon of inversion in your own words.

Part 3: Field of view

Onfield of viewis the amount of sample you see when viewing targets. The field of view decreases at higher magnifications.

Procedure

Place the blue plastic ruler along the stage opening so that the edge of the ruler appears as a vertical line through the diameter of the field. Calculate the field size in millimeters for each objective lens.

Scan __________ Low (10X) __________ High (40X) __________

Part 4: Depth of Field

Ondepth of fieldis the thickness of the sample that remains focused at a given magnification. The depth of field decreases at higher magnifications.

Procedure

Get a slide of colored threads and view it scanned or at low power. Then determine what color the thread is:

• On the top
• In the middle
• W tle

Tip: Following the focusing instructions, focus on the area where the three threads intersect. Use fine focus to recognize the order of the threads.

Part 5: Making a wet fix

Throughout the semester, you are expected to successfully complete a series of simple slide preparations, known as "wet montages". The sample to be observed is placed on a clean glass slide, a drop or two of water is added and a coverslip is carefully placed over the water and sample. Your instructor will demonstrate this technique to you.

Prepare a wet amount of pond water by following the procedure below:

1. Using a disposable plastic pipette, place a drop of pond water on a clean glass slide.
2. Cover the pond water on the slide with a coverslip. Try to place the slide at an angle to avoid air bubbles.
3. Wipe excess liquid from the slide.
4. Place the slide under the microscope to view the sample.
   1. Start with the purpose of the scan
   2. Carefully switch between low and high goals as needed.

We hope you will see living organisms in your pond water. If you see green material, it's probably some type of plant. See if anything moves. In the space below, describe what you see under the microscope and make a simple drawing.

Part 6: Stereo microscope

The instructor will demonstrate the correct use of the dissection scope. These microscopes typically offer lower magnification than the compound microscope you are using.

Procedure

View available samples on the lab bench with a dissecting microscope. Notice that the microscope has two light sources, one from below and one from above. Samples are not always mounted on a slide. It should also be remembered that objects viewed under a dissecting microscope are three-dimensional.

Microscope cleaning and care

1. Describe how and with what you will clean the microscope lens.
2. List four things you should do after using your microscope after you finish your lab.