Microscope Labbiologymr. Banerjee

Microscope LabBiologyMr. Banerjee

Microscope Lab

This lab will have you in teams. Each team must submit a lab write up. The lab write up for this activity will be handwritten and in a duo tang. One person will need to read the directions at a lab station while the other manipulates the microscope. It is essential that all team members become familiar with the appropriate and efficient use of the compound light microscope so make sure all of you use the microscope as instructed.

The most widely used scientific instrument in Biology is the Microscope. Without the microscope, an understanding of the basic nature of life would not be possible. There are several types of microscopes. Some use light to establish an image of a subject and others use electrons. We will be using compound light microscopes to look at subjects that are 5mm or smaller. We will use stereo microscopes (dissecting microscopes) to look at specimens larger than 5mm.

The compound light microscopes we will be using have three objective lenses. The shortest has a magnification of 4x (scanning power), the middle sized one has a magnification of 10x (low power), and the largest is 40x (high power). Look at the number on the lens to determine which you have.

Materials:

compound light microscope
stereo microscope
lens paper
microscope slide
microscope cover slip / slide with the letter e (Students prepare)
slide with crossed threads (Students prepare)
slide with yeast organism (Students prepare)
(Prepared slides are expensive and very delicate - be very carefully not to drop these slides: you drop it...... you buy it!)

Yeast Preparation:

Yeasts are unicellular fungi that are used to make alcoholic beverages such as wine and beer, and also in baking. They consume sugars and, through the process of fermentation, produce alcohol and carbon dioxide.

Materials:

Microscope LabBiologyMr. Banerjee

· Compound microscope

· One packet of active yeast

· Dropper or pipette

· 1 cup warm water

· Microscope slides and cover slips

· 1 tablespoon of sugar

Microscope LabBiologyMr. Banerjee

Instructions:

To begin, you need to prepare a yeast solution. Add the yeast and sugar to the warm water. Stir and let it sit for 45 minutes. Once the yeast solution is ready, place a small drop onto a slide and cover with a coverslip. Observe the yeast cells under high power. You may see some cells in the process of reproducing. In yeast, this is known as budding.

Procedure Part A:

It is essential that you clean up your lab station thoroughly at the completion of the lab activity. The cord of your microscope must be wrapped around the base (for storage and to avoid tripping over cord) of the microscope and the plastic bag placed back over top (to keep dust from settling on the lenses) when done. When moving the microscope to a new location ALWAYS hold one hand under the base and the other should be firmly grasping the arm. You drop the microscope.... you bought it!!!

Using the diagram below become familiar with ALL THE PARTS OF THE MICROSCOPE and be able to discuss their functions.(Knowing the functions of the parts of a microscope will be essential for successful usage of the scope and performance on an upcoming test)

1. Ocular Lens (eye piece) - magnifies image

2. Nose Piece (turret) - allows user to rotate preferred objective lens into position

3. Objective Lenses - allows user to have multiple levels of magnification

4. Stage - specimen to be observed is placed on here

5. Diaphragm - regulates amount of light passing through stage

6. Light Source - if you can't figure out the function of this you should not be in this class

7. Arm - one hand needs to firmly grasp this when moving a microscope

8. Stage Clips - these hold the slide in place on the stage (we will rarely use these)

9. Coarse Adjustment - used to move the stage up or down in large increments

10. Fine Adjustment - used to move the stage up or down in extremely small increments (fine focus)

11. Light Switch -

12. Base - one hand needs to be under this when moving microscope

Questions:

1- If total magnification is determined by Multiplying the ocular lens magnification by the objective lens magnification, what three different levels of magnification can be achieved by your microscope?

2- The thickness and transparency of the specimen being viewed under the microscope can vary dramatically (ie: fly wing, dark green leaf).

A) What microscope part, which should be adjusted every time a specimen is placed on the stage, would you use to establish ideal illumination?

B) Which setting might you use for the fly wing?

Procedure Part B:

· Place the microscope so that the arm faces you and the base is approximately 5cm from the edge of the lab bench.

· Clean the eyepiece and the objective lenses with lens paper only. Never use paper towels or your shirt both of which may scratch the lens. Use a gentle circular motion when cleaning a lens.

· Plug in the microscope and turn on the light.

· Open the diaphragm to the largest opening directly beneath the opening in the stage.

· Turn the nosepiece so that the scanning power objective lens (smallest objective lens) clicks into place directly above the opening in the stage. You will always start with scanning power to locate the specimen before changing to higher magnifications.

· Use the coarse adjustment knob and lower the stage as far as it will go.

· Locate the slide with the letter e on it and place it on the stage SO THAT IT IS RIGHT SIDE UP AND APPEARS EXACTLY AS YOU WOULD EXPECT TO READ IT!. Move the slide so that the letter e is as close to the middle of the opening in the stage as possible. DO NOT USE THE STAGE CLIPS. Most of our work will require moving the slide on the stage and the clips tend to become more of an inconvenience than help.

· Use the coarse adjustment knob and raise the stage as close to the lens as is possible.

· Use the fine adjustment knob (moving the stage away from the lens) to get the letter e CLEARLY FOCUSED (not blurry at all).

Questions:

3- While looking at the letter e through your microscope, gently move the slide to your left and right and then up and down. A) What happen to the image when you move the slide to the right? B)What happens to the image when you move the slide up?

4- Pay close attention to the orientation of the letter e while observing it through the microscope at scanning power (40x). Using the plastic cup provided trace a small circle on your lab write up and make a simple drawing of the letter e's orientation (how it now looks while under the microscope).Accuracy is critical.

Procedure Part C:

· While looking at the letter e through your microscope gently move the slide so that the letter e is as close to the center of the field of view as is possible.

· Carefully rotate the nosepiece so that the Low Power Objective Lens (middle sized lens) clicks into place directly above the opening in the stage.

· While viewing the letter e pay close attention to how much of the field of view is occupied by the letter.

· Place the plastic ruler on the slide and look carefully at the number of millimeter lines that can be seen in the field of view. The image below gives you an idea of how to determine total diameter of the field of view.

Questions:

5- Knowing the diameter of the field of view at low magnification (100x) estimate the total width of the letter e on your microscope slide.

6- Use the plastic cup to trace a small circle on your lab write up and in the circle draw the letter e as it appears under your microscope at low power (100x)

Procedure Part D:

· Remove the plastic ruler and move the slide so that the letter is exactly in the middle of the field of view.

· Lower the stage by turning the coarse adjustment one or two turns.

· Rotate the High Power objective lens into position.

· While looking at the stage from the side, use the coarse adjustment to move the stage so that the lens is as close to the slide as is possible without touching it.

· Move your eye over the ocular lens and use the fine adjustment to focus the image.

Questions:

7- Use the plastic cup to trace a small circle on your lab write up and in the circle draw the letter e as it appears under your microscope at high power (400x or 430x depending upon microscope)

8- Using the formula shown below estimate the diameter of the field of view at 400x

? / low power field of view
=
low power magnification / high power magnification

9- Why can't we simply place the ruler underneath the microscope at high power to determine field of view?

Procedure Part E:

· Rotate the nose piece so that the scanning power objective lens clicks into place directly over the opening in the stage.

· Prepare a slide with three different coloured threads & cover with a cover slip.

· Place the slide with the crossed threads onto the stage.

· Focus your microscope so that all three threads are clearly visible.

· Move the slide so that where they cross over is in the middle of the field of view.

· Carefully rotate the nose piece so that the low power (100x) is directly over the opening in the stage

· Move the slide so that you can observe where the three threads cross

· Lower the stage by rotating the coarse adjustment one or two turns.

· Rotate the High Power objective lens into position.

· While looking at the stage from the side, use the coarse adjustment to move the stage so that the lens is as close to the slide as is possible without touching it.

· Move your eye over the ocular lens and use the fine adjustment to focus on the area where the three threads cross

Questions:

10- Use the plastic cup to trace circle on your lab write up and in the circle draw the crossed threads as it appears under your microscope at high power (400x or 430x depending upon microscope). Use the coloured pencils that are available to properly represent each of the three threads.

11- Were all three colors clearly focused (all three equally clear at the same time) at scanning power? Low power? High power?

12- Depth of field is the amount of vertical space in focus at a particular magnification. What seems to happen to the depth of field as magnification increases?

13- Working distance is the space between the objective lens and the stage. What seems to happen to working distance as magnification increases? Why should we lower the stage before rotating the nose piece to high magnification?

14- What seems to happen to the diameter of the field of view as magnification increases?

15- Explain why it is important to move the specimen we want to view to the very middle of the field of view before switching to a higher magnification.

Procedure Part F:

Most specimens we will look at this year will require a "Wet Mount". This will involve using a small square plastic cover slip to compress the specimen into a small a vertical space as possible and create a tight seal so that it stays in place on the slide. The next procedure will take you through the basics of how to make a wet mount preparation.

· Acquire a single hair from either you or your partner.

· Place the hair in the middle of a clean blank slide

· Add two drops of water at the center of the slide where the hair lays across it.

· Take the small square "Cover Slip" and let the edge of the slip rest on the slide and lean at a 45 degree angle over the hair where the drops of water were added.

· Allow the cover slip to fall, keeping the one edge in contact with the slide, over the hair and form a tight seal with the two drops of water.

· Use the coarse adjustment knob and lower the stage as far as it will go.

· Place the slide on the stage of your microscope attempting to position the area where the hair is over the middle of the opening in the stage.

· Use the coarse adjustment to focus.

· Carefully view the hair under low power and high power as well.

· Attempt to locate the numerous scales of protein that make up the human hair.

Questions:

16- Knowing the diameter of the field of view at different magnifications, what is the approximate width of your hair?

Procedure Part G:

You will prepare and observe cells from both a plant (onion) and an animal (your own cheek cells!!).

Materials: You will need the following materials: glass slide, coverslip, toothpick, iodine stain, onion tissue, and your own cheek cells.

Procedure:

17. use the plastic cup to trace circle on your lab write up and in the circle draw the prepared and observed samplings of both types of cells as possible and sketch them.

You are also required, on your sketch, to label any visible parts or organelles, as well as list what magnification you used. Lastly, you need to write down any other interesting observations (difference between cells, cool things you saw, etc.).