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Biology 11

Using the compound light microscope

Part A: Handling the microscope:

1. Remove the microscope from its storage area, using both hands: one grasping the arm and the other supporting the base. USE ONLY YOUR ASSIGNED MICROSCOPE!

2. Carry the microscope (with 2 hands) in an upright position (stage must be horizontal).

3. Never touch a lens with your finger, nor remove lens from microscope.

4. To clean, rub lens lightly with lens paper (not paper towel). Do not reuse lens paper.

Part B: Parts of the microscope

Use the microscope diagram below to identify the parts and their function. Refer to pages 21-22, 1088-1089 in your Biology text for help.

| |Name |Function |

|A |Eyepiece (ocular lens) | |

|B |Objective lenses | |

|C |Nosepiece | |

|D |Stage | |

|E |Diaphragm | |

|F |Base | |

|G |Light source | |

|H |Coarse adjustment | |

|I |Fine adjustment | |

|J |Stage adjustment knob |Moves the stage along with the slide |

|K |Stage clips | |

|L |Arm | |

Part C: Microscope calculations

In microscopy, most measurements are given in micrometers, not millimetres. Micro mean one millionth. In other words, 1 000 000µm = 1 m and so 1000µm = 1mm. Using micrometers minimizes the use of decimals. Remember: 1000µm = 1mm 10mm = 1cm

Convert the following:

1. 3000µm = _____________ mm

2. 100µm = ______________ mm

3. 10µm = _______________ mm

4. 0.75mm = _______________ µm

5. 0.03mm = _______________ µm

6. 1.0 cm = _________________µm

Part D: Measuring Microscopic Objects

It is often necessary to know the dimensions of the object being examined under the microscope. The size of a microscopic object can be estimated by comparing it with the size of the circular field of view.

Magnification:

Your microscope has four different objective lenses (low, medium, high, and oil immersion) each with a different magnification. The image is also magnified by the eyepiece.

Field of View:

Use a clear ruler and place it, lined up, on the stage to determine the field of view with each lens.

Low power = mm or (m

Med power = mm or (m

High power = mm or (m

Actual Size of an Object:

It is often necessary to know the dimensions of the object being examined under the microscope. The diameter of a cell (its actual size) can be estimated by comparing it with the size of the circular field of view and the number of cells it would take to fit across the field. See formula below.

To find field of view, use the figures you collected above. To determine the number of cells, you simply count how many fit across the circular field (in some cases you may have to draw the same image across the field of view approximating the size).

Example…

(on low power)

Practice Problems: How long are these objects under the following powers?

a) low power? b) medium power? c) high power?

d) How would you do this differently if I asked how wide the objects are?

Magnification of Drawings

When you DRAW a specimen, you may also be required to show how much bigger your drawing is compared to the actual specimen. This is called the magnification. To accomplish this you must know the actual size of the cell and the drawing size. The actual size can be calculated using the equation above. To determine the drawing size, you will have to measure the drawing using a ruler with mm and then convert to µm.

Example: An object under the microscope is determined to be 250 (m in length (actual size already calculated for you), and a drawing of the object is made in which the drawing is 15 cm in length. Find the magnification:

Problem: Calculate the magnification of an object examined under medium power that covers a third of the field of view, if your drawing measures 12 cm.

Part E: Characteristics of microscope images

1. Prepare a wet mount slide of the letter ‘e’ following your teacher’s instructions.

2. Place the slide on the stage under low power. Ensure that the ‘e’ is in the middle.

3. Using the coarse adjustment knob, move the stage to the very top. While looking through the ocular lens, slowly lower the stage with the coarse adjustment knob until you see the image.

4. When the image is in sight, use the fine adjustment knob to focus.

5. Use the iris diaphragm to adjust the light for a better viewing.

6. Look at the ‘e’ on the slide with unaided eyes and then look at the ‘e’ through the microscope. What do you notice about the ‘e’? _________________________

7. Look through the ocular lens and record what happens to the image when you:

a. Move the slide to the right ___________________

b. Move the slide to the left __________________

c. Move the slide up ____________________

d. Move the slide down ___________________

8. Now, without moving the stage, flip to medium power. Use only the fine adjustment knob to focus. Adjust the diaphragm if necessary.

9. Without moving the stage, flip to high power. Look from the side to make sure the lens and the slide don’t hit one another. Use only the fine adjustment knob to focus. Adjust the diaphragm if necessary.

Note: If you lose sight of your image, you must return to low power and start from the beginning.

Part F: Depth of field

1. Prepare a wet mount of two strands of different colour hair. Make a cross with the two strands and place the cover slip over the cross.

2. Place the slide on the stage and ensure the crosses of hairs are in the center.

3. Focus on low power and draw what you see. Remember to use the same technique used with the letter ‘e’ to rotate to higher objective lenses.

4. Focus on medium power and then high power. You will notice that it is not possible to get both hairs in focus at the same time. This is because the field of view is three-dimensional with depth as well as width. Draw two different diagrams, each focusing on different strands on high power.

Strand 1 Strand 2

Fill in the missing blanks and calculate Actual Size & Magnification for each of the following:

Formulas (look in your notes):

Actual Size = Magnification =

|Lens Power |Field of View |# of cells across |Actual Size |Drawing Size |Magnification |

| | |f.o.v. | | | |

|High | |90 | |9.3 cm | |

|Medium | |3 | |12.4 cm | |

|Low | |15 | |60 mm | |

| |1800 um |20 | |85000 um | |

| |450 um |2 | |7 cm | |

| |4500 um |5 | |14 cm | |

|Low | |2 | |8.8 cm | |

| |1800 um |6 | |66 mm | |

|High | |10 | |10 cm | |

| |450 um |45 | |4.5 cm | |

-----------------------

Actual size = Field of view

(diameter) Number of cells

Magnification = Drawing size

Actual size

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