Microscope 55

Biology Microscope Laboratory • Be able to identify the structures and their functions. • Know and be able to use the vocabulary of the microscope. • Be able to focus properly on a slide on high power in two minutes. • Be able to guestimate the size of a cell. • Be able to tell the top from bottom colored thread.

Microscope History We will not test on this material, but it may help you understand where microscopes came from, and how they came to have so many parts.

• History of the Light Microscope A good history in six chapters • Microscope A-Z A very short history • A recreation of the simple, single-lens microsco microscopists saw.

Choose the right microscope for the job Dissecting Scope

larger working distance

less magnification

Compound Light Microscope

smaller working distance

more magnification

Links to other Microscope Pages Different kinds of microscopes • Fluorescence Microscope How does it work? • The Scanning Electron Microscope(SEM) is one of the most versatile and widely used tools of modern science as it allows the study of both morphology and composition of biological and physical materials. • STM Project Scanning Tunneling Microscope. This webpage covers the characteristics of an electron microscope and of an optical microscope, versus an STM. A bit technical.

Get your microscopes in H-325 • Carry a dissecting scope with two hands to your lab desk • turn the light on • practice focusing on your fingernails or jewelry

• Carry a compound light microscope with two hands to your lab desk • be sure the base is at least a thumbs length in from the edge of the desk. • Find the following structures on the scope

Be able to identify the structures and their functions • • • •

Ocular Body tube Revolving nosepiece Objectives – – – –

Scanning objective Low power objective High power objective Oil immersion objective

• Eyepiece - 10X • light tight tube • change objectives • second set of lens – – – –

4X 10X 40 or 43X 97 or 100X

Be able to identify the structures and their functions • Arm • Base • Stage • Stage clips / mechanical stage • Aperture

• connects body to base • firm foundation for scope • Platform for slides • holds slide / moves slide • hole in stage for light

Be able to identify the structures and their functions • Condenser • Iris diaphragm • Coarse focus • Fine focus • Light

• focuses light on slide • adjusts amount of light • focus on scanning and low • focus on high and oil • illuminate the slide

Links to other Microscope Pages • Microscope parts page Here is another school’s look at some parts of the microscope Microscope laboratory Still another school’s look at some parts of the microscope in their microscope lab.

Know and be able to use the vocabulary of the microscope. • Magnification • Resolution • Numerical aperture

• image larger • image clearer • ability of lens to gather light

Know and be able to use the vocabulary of the microscope. • Working distance • between lens & stage • Depth of field • bottom to top of slide • Parfocality • all objectives in reasonable focus at the same time

Links to other Microscope Pages • Depth of field discussion

• Resolution discussion

Total magnification ocular power multiplied by objective power • • • •

Ocular = 10x scanning = 4x low = 10x high = 40x or 43x

• oil

= 97x or 100x

• • • • • • •

Total magnification 10 x 4 = 40 10 x 10 = 100 10 x 40 = 400 or 10 x 43 = 430 10 x 97 = 970 or 10 x 100 = 1000

Be able to focus properly on a slide on high power in two minutes. We call this the “two minute drill”

Clean the microscope with lens tissue only! only!

Turn on the light and adjust it to the lowest amount of light

There will be a eight step summary slide at the end of this section

Start with the lowest power objective in place.

Place the precleaned letter e slide on the stage with the specimen in the light from the condenser.

Bring the stage and the lowest power objective as close together as you can. You should hit the automatic stop.

Looking in the ocular, slowly focus away with the coarse focus until you see something, even dirt on the slide. If you need more light, open the iris diaphragm a bit.

Move the slide slowly until you have centered the object you are looking at. Focus with the coarse focus until it is in the best possible focus.

Turn the revolving nosepiece to the low power objective. Be sure to click it in place. Center the slide. Focus with the coarse focus until it is in the best possible focus.

Turn the revolving nosepiece to the high power objective. Be sure to click it in place. Center the slide. Focus with the fine focus until it is in the best possible focus.

Oil immersion lens When magnification is this high, any loss of light will degrade the resolution, making the image fuzzier and harder to see. A small drop of oil, between the slide and the objective lens, will prevent light from scattering and preserve the image.

To focus on oil, first make sure you have focused as well as you can on high power • Move the revolving nosepiece until you are halfway between high and oil. • Place a small drop of oil on the slide where the lens will be. • Remember, the more oil you put on, the more there is to clean off later. • Move the oil objective in place and focus with the fine focus only!

Be sure to clean the lens and the slide with with lens lens tissue tissue when when you you slide are done done are The oil will seep into the microscope and loosen the cement, as well as dry to a scummy mess on the lens.

Focusing summary 1 2 3 4 5 6 7

Clean the microscope with lens tissue start on the shortest objective turn light down and center slide over light bring objective and slide together until they stop use coarse focus on scanning, turn to low power use coarse focus on low, turn to high power use fine focus on high, add a drop of oil and turn to oil immersion objective 8 use fine focus on oil ✳ adjust light light at at each step as needed

Links to other Microscope Pages • Microscope trouble shooting chart This may help you if you are having trouble finding or focusing on your slide.

Guestimate the size of a cell • Field of view is the white circle of light you see when you look in the ocular. • If you know the diameter of the field of view, you can guestimate the size of a cell • count how many cells could fit across the field of view, and divide the field of view by that number. • We measure microscopic things in micrometers, or um.

Guestimate the size of a cell • Light microscope measurements are in micrometers, or um. • There are 1,000 um in one millimeter, or mm. • One mm is about as thick as a fingernail. • There are 10 mm in one centimeter, or cm. • There are about 2 1/2 cm in one inch.

Guestimate the size of a cell • The field of view is the circle of white light you see when you look through the ocular • This can be measured with a microscopic ruler or micrometer. • It will look something like this under the scanning objective.

2 mm or 2000 um

Guestimate the size of an object • The scanning power field of view is about 4500 um • How many stars would it take to cover the diameter of the field of view? • Seven stars would do it. • Divide seven into 4500 um • the star is about 630 um • an answer between 560 + 640 um is reasonable

Guestimate the size of an object • The low power field of view is about 1700 um • How many wood chips would it take to cover the diameter of the field of view? • Four chips would do it. • Divide four into 1700 um • the chip is about 425 um

Guestimate the size of a cell • The high power field of view is about 450 um • How many cells would it take to cover the diameter of the field of view? • About 14 or 16 cells would do it. • Divide 14, 15, or 16 into 450 um • The cell is about 28-32 um

Guestimate the size of a cell • The oil immersion field of view is about 170 um • How many cells would it take to cover the diameter of the field of view? • Five cells would do it. • Divide five into 170 um • The cell is about 34 um

Depth of field • Microscopes “fool the eye”. • What ever is in focus looks like it is on top. • To tell top from bottom, you must focus through the layers. • We will do this with a colored thread slide.

Depth of field • Colored thread slides Colored threads • red, blue + yellow threads in all Back arrow 4 times combinations to repeat • focus away from the slide • bottom thread will be label in focus first • keep keep light level level low

Low power objective

coverslip threads slide Focal point

Links to other Microscope Pages • Nanoworld HomePage Try the image gallery and the Transmission Electron Microscopy page. The “What is it?” quiz is fun.

Lab Exam - 10 Questions • Be able to identify the structures and their functions. • Know and be able to use the vocabulary of the microscope. • Be able to focus properly on a slide on high power in two minutes. • Be able to tell the top from bottom colored thread. • Be able to guestimate the size of a cell.

Lab Exam • The Lab Exam questions will be similar to the following practice questions. • These questions are examples only, and do not cover all possible questions that could be asked. • Where possible, a short answer will appear on the slide.

Identify the structure and it’s function 1. • Ocular • Body tube 2.

• Eyepiece - 10X • light tight tube

• Revolving 3. nosepiece

• change objectives

Identify the structure and it’s function • Arm

4.

connects body to base

• Base

5.

firm foundation for scope

• Stage 6. clip / mechanical stage • Stage 7.

holds slide / moves slide

Platform for slides

Identify the structure and it’s function Pointing to lever underneath

• adjusts amount of light • focus on scanning

• Iris diaphragm

• Coarse focus

• Fine focus 8.

and low • focus on high and oil

Define the following terms • 11. Magnification • 12. Resolution • 13. Numerical aperture

• image larger • image clearer • ability of lens to gather light

Define the following terms • 14. Working distance • between lens & stage • 15. Depth of field • bottom to top of slide • 16. Parfocality • all objectives in reasonable focus at the same time

What is the total magnification for these objectives? Total magnification

• 17. scanning = • 18. low = • 19. high = • 20. oil

=

• • • • • •

10 x 4 = 40 X 10 x 10 = 100 X 10 x 40 = 400 X or 10 x 43 = 430 X 10 x 97 = 970 X or 10 x 100 = 1000 X

Ocular X objective

Find an Instructor or Lab Technician and ask them to watch you focus properly on a cheek cell slide on high power in two minutes.

You should also be able to describe the proper procedure for focusing in a short essay. The focusing summary slide is a good review.

After you have focused on the cheek cell slide, guestimate the size of the cheek cell and ask the instructor to check it.

Find an Instructor or Lab Technician and ask them to watch you focus properly on a blood slide on oil immersion.

After you have focused on the blood cell slide, guestimate the size of the blood cell and ask the instructor to check it.

Focus properly on a colored thread slide and determine which thread is on the bottom and which thread is on the top. Check the label on the slide for the correct order.

These questions are similar to the questions you will answer on the laboratory practical. Good Luck!

The End