Embedded Files
Skill 4: Create a wet mount slide. Find and focus the specimen at all magnifications using proper procedures.
Skill 4: Create a wet mount slide. Find and focus the specimen at all magnifications using proper procedures.
Microscope Basics
Microscope Basics
Read and understand the following:
Read and understand the following:
- Your microscope is in the end cabinet at your lab station.
- Your microscope is in the end cabinet at your lab station.
- Use it with the cord facing your body.
- Use it with the cord facing your body.
- When raising the stage, take your eye away from the ocular lens, or eyepiece, and look at the stage to ensure that it does not hit the slide.
- When raising the stage, take your eye away from the ocular lens, or eyepiece, and look at the stage to ensure that it does not hit the slide.
- The shortest lens is the ‘scanning power’ and is used for finding the object on the slide. It has a wide field of vision, but not much magnification. This lens is red on our microscopes. "Scanning is short."
- The shortest lens is the ‘scanning power’ and is used for finding the object on the slide. It has a wide field of vision, but not much magnification. This lens is red on our microscopes. "Scanning is short."
- The medium length lens is the ‘low power.’ It has a higher magnification, but a smaller field of vision. "Yellow is low."
- The medium length lens is the ‘low power.’ It has a higher magnification, but a smaller field of vision. "Yellow is low."
- The longest lens is the ‘high power.’ It has the highest magnification, but the smallest field of vision. High power is blue on our microscopes.
- The longest lens is the ‘high power.’ It has the highest magnification, but the smallest field of vision. High power is blue on our microscopes.
- Remove and clean slides before putting away microscope.
- Remove and clean slides before putting away microscope.
- Unplug the microscope by the plug, not the cord. Do not wrap the cord around the cord keeper.
- Unplug the microscope by the plug, not the cord. Do not wrap the cord around the cord keeper.
- Lower the stage. Switch to the scanning lens. Always put your microscope back where you found it.
- Lower the stage. Switch to the scanning lens. Always put your microscope back where you found it.
- Ask me to check your lab station before leaving .
- Ask me to check your lab station before leaving .
Quiz your partner(s) and have them quiz you before moving on.
Quiz your partner(s) and have them quiz you before moving on.
When looking through a microscope eyepiece, keep both eyes open to reduce eye strain and headaches. If you determine which eye is dominant and look through that eye, it is easier to focus on the slide and ‘tune out’ what your non-dominant eye is seeing.
When looking through a microscope eyepiece, keep both eyes open to reduce eye strain and headaches. If you determine which eye is dominant and look through that eye, it is easier to focus on the slide and ‘tune out’ what your non-dominant eye is seeing.
Make a triangle with your hands as shown.
Make a triangle with your hands as shown.
Frame the American flag at the front of the room with the triangle.
Frame the American flag at the front of the room with the triangle.
Close your left eye. If you still see the flag, your right eye is dominant. If the flag disappears, your left eye is dominant.
Close your left eye. If you still see the flag, your right eye is dominant. If the flag disappears, your left eye is dominant.
Pick up a worksheet from the conference table to write your lab answers on.
Pick up a worksheet from the conference table to write your lab answers on.
Objective 1: Handling the microscope
Objective 1: Handling the microscope
Work your way through the lab, following all instructions, until you reach a number. Read the question and then write the answer on your answer sheet. DO NOT work from the lab sheet or you will miss most of the information.
To carry: grasp the arm with one hand and the base with the other. Use two hands and hold the microscope close to your body.
1. What are the two parts of the microscope that you hold on to when carrying the microscope?
Practice using the coarse adjustment knob without a slide on the stage. Turn the knob toward you and notice which way the stage moves. Try turning it away from you and note the stage direction. Remember that you look at the stage, not through the eyepiece, when raising the stage with the coarse adjustment knob.
2. When you turn the knob toward you (counter-clockwise when looking at the right knob), which way does the stage move? When you turn it away from you (clockwise when looking at the right knob), which way does the stage move?
When preparing to focus on a slide, remember to always start with your scanning objective lens (the shortest one). Raise the stage as high as you can, while looking at the stage to ensure that you don’t hit the objective lens with the slide. Once you have raised the stage, make sure you know which way to turn the knob to lower it. Begin to slowly lower the stage while looking through the eyepiece, keeping both eyes open. Once the object is in focus, switch to the next higher power objective lens, without adjusting the focus. When switching from a lower power lens to a higher power one, look at the lens from the side to ensure that you don’t hit the slide with the lens. When you are not using the scanning lens, you only use the small, fine adjustment knob.
3. Which objective lens do you always start with when examining a specimen?
Objective 2: Preparing a wet mount slide
Objective 2: Preparing a wet mount slide
Handle slides and cover slips by the edges to avoid fingerprints.
Cut one lowercase ‘e’ out of the newspaper.
Place a drop of water in the center of the slide and gently set your "e" on top. Hold the cover slip at a 45-degree angle over the drop of water and touch the slide. Drag the cover slip toward the water until it just touches. Water will start to climb the cover slip. Let go of the cover slip and let it drop onto the slide. By touching the water and then dropping the cover slip, it pushes air out in front of it and minimizes trapped air bubbles.
Review: Which property of water causes the water in the drop to stick to the cover slip?
If you need more light or it is too bright, adjust the diaphragm, found under the stage. The larger holes let more light through. Find the diaphragm, even if you don't need to adjust it. Look at the bottom of the stage from underneath. You will get the best resolution with the least amount of light.
Lower the stage all the way. Put your wet mount slide onto the microscope’s stage and use the stage clips on the edges of the slide to secure it in place. Turn the nosepiece so that the scanning objective lens is in place. Raise the stage as high as you can without hitting the slide on the objective lens. Remember to look at the stage when raising it, not through the eyepiece, or ocular lens. Focus under scanning power by slowly lowering the stage while looking through the eyepiece, keeping both eyes open. When it comes into focus, use the fine adjustment knob to "fine tune" your focus. Everyone's eyes are different, so you may have to adjust the fine focus when switching between partners. When switching from a lower power lens to a higher power one, look at the lens from the side to ensure that you don’t hit the slide with the lens. You should be able to clearly see the letter “e” on the slide. Be sure it is centered in your field of vision.
Ask me to look at your slide on scanning power before moving on.
Ask me to look at your slide on scanning power before moving on.
When switching to the low power objective, do not change the focus knobs. Turn the low power objective into place (yellow is low). The image should almost be in focus already. Use only the fine focus when looking through the eyepiece. Move the slide around and focus to see small irregularities in the “e”.
NOTE: slides are easily broken and lenses damaged by raising the stage while looking through the eyepiece. Always look at the stage when using the gross adjustment (big knob) to raise it, to avoid hitting the slide.
Which magnification, scanning or low, is more appropriate for looking at the “e”?
How does the “e” appear to be positioned compared to the way it looks on the stage?
Move your slide to the left. Which way does the image of the “e” appear to move?
Move your slide toward you. Which way does the image of the “e” appear to move?
What can you conclude about the way the image moves as compared to the way you are actually moving the slide?
Throw the letter "e" in the trash can. Rinse the cover slip and place it on the towel by the sink in the corner of the room. Continue to use the same slide after you dry it with lens paper. The lens paper is by the corner sink. Do not throw it away. Just spread it out to dry for the next person.
Objective 3: Depth of field
Objective 3: Depth of field
Under the compound light microscope most objects are observed in a drop of water. If you think of that drop of water as a pond and the microorganisms as fish in the pond you will begin to see why it is important to be able to focus at different depths. This is similar to taking a picture with your phone. You have to choose whether to focus on someone in the foreground or something in the background. Depth-of-field focusing is always done with the fine adjustment knob. You will not use water or a coverslip for this part of the lab.
Lay a piece of white thread on your slide. Lay a red piece of thread across it, making an ‘x.’
Find the place where the two threads cross under scanning power and bring it into focus. Adjust the slide so that the point at which they cross is in the center of your field of vision.
1. Can you see both threads in focus at the same time?
Switch to low power and observe the point at which the threads cross.
2. Can you see both threads in focus at the same time with this power?
Slowly turn the fine adjustment up and down until you have seen each of the threads in focus. Draw your observation (in pencil) of the thread on scanning power and low power in the circles on your answer sheet. Color the red thread and label the thread which is on top. Be sure to indicate the magnification in the lower right hand corner of your drawing. (ex. X40)
The magnification of your microscope is the product of the separate magnifications of the eyepiece and the objective lens. The eyepiece, or ocular lens, is 10x and the low power objective is 10x, therefore the magnification under low power is 100x. The equation for total microscope magnification in found below:
Total magnification = (eyepiece magnification) x (objective magnification)
Objective lenses on this microscope
Scanning objective = 4x
Low power objective = 10x
High power objective = 40x
Return the threads to the petri dish. Do not throw them away. Remove your slide from the stage. Rinse it, dry it with lens paper, and put it on the towel by the sink in the corner.
Objective 4: Microscopic measurements
Objective 4: Microscopic measurements
The purpose of a microscope is to magnify your specimen. Microscopes use two lenses – the eyepiece and an objective – to magnify the image. The magnification number indicates how many times larger the object appears than its actual size. If the magnification of an object is 10x, it will appear 10 times larger than it really is.
The magnification of your microscope is the product of the separate magnifications of the eyepiece and the objective lens. The eyepiece, or ocular lens, is 10x and the low power objective is 10x, therefore the magnification under low power is 100x. The equation for total microscope magnification in found below:
Total magnification = (eyepiece magnification) x (objective magnification)
Objective lenses on this microscope
Scanning objective = 4x
Low power objective = 10x
High power objective = 40x
1. What is the total magnification using the scanning power lens on your microscope? Total mag on low? Total mag on high?
To measure the field of view of a microscope you must use a unit called a micrometer, or micron (µm). A micrometer equals 0.001 mm.
2. How many micrometers are there in one mm?
3. Is it possible to divide a millimeter into that many micrometers using just your eyes? Why or why not?
Place the millimeter section of a plastic ruler over the center of the stage opening of the microscope. Using the low power lens, locate the measured lines of the ruler in the center of the field of view. Move the ruler so that one of the millimeter lines is visible at the left edge of the field of view, as shown.
4. Remember that the distance between two black lines is one millimeter. Estimate the diameter (in mm) of the field of view on low power.
5. Calculate the diameter of the low power field of vision in micrometers.
Cleanup - Use the lens paper by the sink in the corner to wipe all four lenses on your microscope. Do not throw away the lens paper. Spread it out to dry and put it back by the sink. Unplug your microscope by the plug. Do not wrap the cord up. Wash your hands with soap and water as we do after each lab. Have your lab station checked before leaving.
Optional - this will help you learn dimensional analysis, which you will need in chemistry
Because the high power field of view is less than one millimeter, it will be difficult to estimate the diameter by the technique you just used to measure the low power field of vision. To calculate the diameter of the high power field use the following calculation.
Low power objective = 10x
High power objective = 40x
10x (low power objective lens) / 40x (high power objective lens)
10/40 = 1/4
This means that with the high power lens, you can only see ¼ of the area that you can see with the low power lens, but it is four times larger so you can see it in more detail.
6. Using the low power field in micrometers from #5 above, calculate the high power field of vision in micrometers.
The following figure is a picture of cells viewed under a microscope. Assume the figure is being observed under low power.
Measure the diameter of the drawing above in millimeters using a ruler. Now measure the cell with the red arrow. The cell is a rectangle that the arrow is inside of. Clean up your lab station, wash your hands and return to your seats to see if you can figure out the dimensional analysis below.
Low power field of view (µm) Size of cell in mm
Size of circle above in mm X (size of cell in µm)
Use your calculation of diameter to determine the size of one cell in micrometers.
Why is it necessary to make your drawings as accurate as you can?
What is the purpose of indicating the magnification on your drawings?
Come check your lab answers. Ask about any which you got wrong so that you can learn from what you missed. You will not turn in your lab. Labs are practice to learn skills. You will take a short quiz on the material from this learning target.
Come check your lab answers. Ask about any which you got wrong so that you can learn from what you missed. You will not turn in your lab. Labs are practice to learn skills. You will take a short quiz on the material from this learning target.
Page updated
Report abuse