BSC 1005L Lab #2: The Microscope

1. BSC 1005LLab #2: The Microscope The Microscope: Protocol and Usage

2. Lab #2: The Microscope • Purpose of lab • Introduce the microscope • Discuss protocols and how to use the microscope • Materials for microscope lab • Microscope • Lens tissue and cleaner • Permanent slide with letter “e” • Permanent slide with color fibers • Clean microscope slides and cover slips • Eyedroppers and clear plastic ruler

3. The Microscopic World of Cells • Every organism is composed of one or more cells • Cells are like miniature “factories” • Main tools to see cells are microscopes • Light microscope • Visible light projected thru specimen • Glass lenses enlarge image • Magnification: increase in object’s apparent size compared to actual size • Resolving power: ability to show two objects as separate

4. History of the Microscope • Dutch spectacle maker Zacharias Janssen credited with the invention of the microscope in 1590 • The compound microscope uses lenses and light to enlarge the image • Also called an optical or light microscope • Contrast this with an electron microscope • The simplest optical microscope is the magnifying glass • Good to about 10x magnification • The compound microscope has two systems of lenses for greater magnification • (1) the ocular or eyepiece that one looks into • (2) the objective lens, which is the lens closest to the object being examined

6. The Microscopic World of Cells • Resolving Power • Human eye = 1/10 of a millimeter (mm) • Light microscope = 0.2 micrometers (~size of a bacterial cell) • Cells first described by Robert Hooke in 1665 • Cells found in every living organism • Led to “cell theory” by mid-1800s = All living things are comprised of cells • All cells arise from previously existing cells

7. Sketch of Robert Hooke’s Microscope (1665)

8. How a microscope works

9. Advances in Microscopy • Electron microscope (EM) • Use began in 1950s • Uses a beam of electrons to resolve objects • Can distinguish objects as small as 0.2 nanometers (= 1/1000 of a micron) • Scanning electron microscope (SEM) • Used to study detailed architecture of the cell surface • Transmission electron microscope (TEM) • Used to explore internal cell structure

10. Stereo microscope Electron microscope

11. Different microscopic views of Euglena

12. Light micrograph of Euglena

13. SEM of Euglena

14. TEM of Euglena

17. The Microscope (page 29) • Part A – Learning to use the microscope • 1. Make a preliminary study of figure 1 on page 30. Be able to identify the arm, stage, and base. • 2. Obtain a microscope. Carry it to your lab bench with two hands: one on the arm, the other under the base. • 3. With the microscope in front of you, return to figure 1 on page 30. Find all of the parts labeled in drawing on your instrument. • 4. Read all of the explanations for the labels following figure 1 on page 31.

18. Test Your Knowledge of the Microscope

19. Test Your Knowledge of the Microscope Ocular Tube Nosepiece Arm Objective Objective Stage Objective Stage clips Coarse adj. Fine adj. Illuminator Base

20. The Microscope (page 32) • Part A – Learning to use the microscope • 5. Determine the magnification capabilities of your instrument. • Total magnification = magnifying power of eyepiece x magnifying power of objective • Eyepiece on our microscope is 10x • Objectives are 4x, 10x, or 40x • Complete the table on page 32 • Part B – Use the microscope with a slide • 1. Obtain a prepared slide containing the letter “e”. Start with the shortest objective (4x) and place the slide in the stage holder.

21. The Microscope (page 29) • Part B – Use the microscope with a slide • 1. Look through the eyepiece after centering the slide. • 2. Practice moving the slide on the stage. • What do you notice about the direction of movement of the image? • 3. Compare working distances and sizes of fields of view. • Working distance = space between the objective and the slide. • Field of view = amount of slide visible at one time. • Why is it easier to find an object on the slide with the 4x objective? • Give two reasons why we start with the 4x objective.

22. The Microscope (pages 34-35) • Part B – Use the microscope with a slide • 4. Experiment with the higher powered objectives • Look at the slide with the “e” using the 10x objective. • What happens to your field of view? • What happens to your working distance? • Repeat using the 40x objective. • Part C – Measuring with a microscope • 1. Find the diameter of the field of view. • Obtain millimeter ruler and use the 4x objective • Place the edge of the ruler across the center of the hole in the stage as shown in Figure 3

23. The Microscope (pages 34-35) • Part C – Measuring with a microscope • 1. Find the diameter of the field of view. • Estimate the diameter of the field of view on low power (4x) • Following the same procedure with the 10x objective • Compare your answers with the numbers in Figure 2 on page 33 • 2. Introduction to the micrometer • = 1/1000 of a mm or 1/1,000,000 of a meter • Express field of view diameter of your lenses in micrometers

24. The Microscope (pages 34-35) • Part D – Depth of Field • Depth of field = amount of vertical space on the slide in focus at any given moment • Gets smaller with increased magnification • Obtain a permanent slide with cross-colored fibers • Examine the slide with the 4x objective • Can you determine which fiber is on top? • Repeat using the 10x objective • Can you determine which fiber is on top? • Repeat using the 40x objective • Which fiber is on top? • Which fiber is in the middle? • Which fiber is on the bottom?

25. The Microscope (pages 37-38) • Part E – Another look at magnification • Eyepiece projects an image that is focused on your retina • Your brain interprets that image • Keep both eyes open when looking into the microscope. • Part F – Look at some other objects • 1. Obtain a blank microscope slide and a cover slip. • Tear off a small piece of note paper. Have one ragged edge and one smooth edge.

26. The Microscope (page 38) • Part F – Look at some other objects • 2. Place a few salt crystals in the center of a dry slide. • Examine with the 4x and 10x objectives. • Are the crystals uniform in size and shape?

27. Exercises – The Microscope • 1. Another name for the eyepiece is _____? (ocular) • 2. The moveable part of the microscope on which objectives are mounted is the _____? (nosepiece) • 3. The total magnification of your microscope when the low power is locked in place is _____? (40x) • 4. If you moved the slide with the letter “e” to the right and away from you, the image moved _____? (left and towards you)

28. Exercises – The Microscope • 5. Whenever you examine a slide, you always begin with the _____ power lens. (lowest) • 6. At what magnification do you see the largest area from your slide? (4x, lowest) • 7. Assume that your instructor have you a 15x ocular that fit your microscope. What would be the total magnification if it were used with the third longest objective? (15 x 40 = 600 times)

29. Exercises – The Microscope • 8. The diameter of the field of view of your slide is approximately _____ when the 40x objective is in use. (0.4 mm) • 9. If after a 10x is carefully focused, the 40x lens is swung into place and it is also in focus with only minor adjustments, the two lenses are considered _____. (parfocal) • 10. Which objective should be in place when the microscope is put away for the day? (lowest, 4x)