BSC 1005LLab #2: The Microscope The Microscope: Protocol and Usage
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
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
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
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
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
Stereo microscope Electron microscope
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.
Test Your Knowledge of the Microscope Ocular Tube Nosepiece Arm Objective Objective Stage Objective Stage clips Coarse adj. Fine adj. Illuminator Base
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.
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.
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
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
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?
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.
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?
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)
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)
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)