The Microscope

For Microbiology Students

 

In this lab the student will learn the following…

1.    Safety Rules

2.    The parts of the microscope

3.    How light travels through the microscope

4.    How to view a slide through the microscope

5.    To define the following terms…

a.    Total magnification

b.    Resolution

c.    Parfocalized

d.    Field of view

e.    Depth of field

6.    The student will also answer the following questions.

a.    Why are we limited to 1,000X magnification in lab?

b.    What is the relationship between light and resolution?

c.    What happens to the amount of light coming through the slide as you increase magnification?

d.    How can you increase resolution when you change to a more powerful objective?

 

I.  Safety Rules

 

1.  When transporting the microscope, hold it in an upright position with one hand

on its arm & the other supporting its base.  Avoid jarring the instrument when

setting it down.

 

2.  Use only special grit-free lens paper to clean the lenses.  Clean all lenses

before & after use.

 

3.  Always begin the focusing process with the low power objective lens in

position, changing to the higher-power lenses as necessary.

 

4.  When using the microscopes, try to keep both eyes open to avoid getting a

headache.

 

5.  Use the coarse adjustment knob only with the low power objective lens.

 

6.  Before leaving lab, remove any microscope slides from the stage, clean the

oil immersion lens with lens paper, rotate the lowest-power objective lens into

position, & cover the microscope.

 

7.  Never remove any parts from the microscope; inform your instructor of any

mechanical problems that arise.

 

8.  Never yank on the electric cord of the microscope.  Always pull it out of the

plug by the base of the cord.

 

II.  The parts of the microscope

Utilizing the microscopes in lab and the diagram in your lab manual, identify the

following microscope parts.

rheostat acts as a dimmer switch for the microscope.  Please

set this dial to 7 if present.

lamp that is in turn focused through the specimen and magnified

by one of the objective lenses.  The condenser optimizes brightness,

evenness of illumination, and contrast.

100x (oil immersion)

Microscope Test

III.  Following light through the microscope

Turn the microscope’s light on.  Follow the path of light through the microscope. 

Using the terms learned above, complete the following list…

 

Light leaves the lamp and follows the following path through the microscope…

  1.  
  2.  
  3.  
  4.  
  5.  
  6.  
  7.  

 

IV.  Magnification and Resolution

 

In a compound microscope, magnification is achieved through the interplay of

two lenses, the ______________________ lens and the

_____________________________ lens.

 

The _________________________________________ of the specimen is equal

to the power of the ocular lens multiplied by the power of the objective lens used. 

The ocular lenses in our lab have a 10x power.

scanning = (10x)(4x) = _____________

low power = (10x)(10x) = ___________

high power = (10x)(40x) = ___________

            oil immersion = (10X)(100X) = ___________

 

Why don't most microscopes magnify more than 1,000x?

Although the level of magnification in a compound microscope is almost

limitless, resolution is not.  

The ____________________________________________ (or resolving

power) is the ability to distinguish that two objects, which are very close

together, are, in fact, two separate objects.

Remember what happens when you enlarge a blurry photo, the mistake

just gets larger.

What affects resolution?

The amount & physical properties of the visible light that enters the

microscope.

In general, as light increases resolution _____________________. 

 

V.  Viewing a prepared slide – the letter e

 

Steps to viewing the slide

 

1.  Rotate the revolving nosepiece so that the scanning objective is in place.  Use

the coarse adjustment knob to move the objective lens and the stage as far apart

as possible.  Turn on the lamp of your microscope.

 

2.  Put the slide on the stage, secure it with the stage clips.  Center the letter e in

the light beam passing through the stage.

 

3.  Use the coarse adjustment knob to bring the objective lens & stage as close

together as possible.

 

4.  Look through the ocular lens & use the coarse adjustment knob to focus

slowly away from the e until it is as clearly focused as possible.  Complete the

focusing with the fine adjustment knob.

Look at the letter e on your stage and then look at the letter e through the

ocular.  Does the letter e appear any differently through the ocular?  If so,

how? ______________________________________________________

Moving the slide

When you move the slide to the right, how does the image appear to

move?  ____________________________________________________

When you move the slide up, how does the image appear to move? 

___________________________________________________________

Changing objectives

Without touching the focusing knobs, increase the magnification by

rotating to the next higher magnification lens, the low power lens.

Why did the image stay in focus, or nearly so?  Because the

microscopes are ______________________________________

**Use only the fine adjustment knob to sharpen the focus.  Using

the coarse adjustment knob could drive the objective lens through

the slide, breaking the slide & possibly damaging the lens.**

What happened to my letter? 

Notice that you can now see only part of the letter e.  Now try looking at

the slide with the high power objective.  Notice you see even less of the

letter e.

The actual area of the slide that you can see through the ocular is called

the ____________________________________________. 

As you increase magnification, you __________________the field of view.

How does this affect my light?

Go back to the scanning objective.  Use the substage diaphragm to adjust

the light to a level that is comfortable to your eyes.

Start with the scanning objective, then the low power, and then the high

power objective.  Note what happens to the light.

As you increase magnification, you ____________________ the amount

of light reaching the ocular. 

This happens because the field of view decreases as the magnification

increases.  The larger the field of view, the more light can pass through. 

As the field decreases (with increasing magnification), less light can pass

through.

How does this affect resolution?

Remember that resolution decreases as light decreases.  Therefore, what

should you do as you increase magnification to increase resolution?

___________________________________________________________

 

VI.  Perceiving Depth with the cross thread slide

The vertical distance that is in focus in the slide is the _____________________.

In this slide, 3 silk threads (a red, yellow, & blue one) are stacked, one on top of

another one.

Start with the scanning objective.  Locate the point where the 3 threads cross

each other.  Notice that some of the fibers of the threads are in perfect focus

while others are slightly blurry.

Change to the low power objective and then the high power objective. 

Compare the scanning objective to the high power objective.  Are more or

less fibers in focus at once with the high power objective?  ___________

 

As you increase magnification, you _________________ the depth of field

 

Return to the high power objective.  Use the substage revolving diaphragm to

decrease the light as much as possible.  This will increase contrast.  Use the

fine adjustment knob to move the objective until the threads are slightly out of

focus.  Then, begin moving the turning the fine adjustment know slowly the other

way, noting which thread comes into focus first, second, and then last. 

 

Which thread do you think is in the middle?_____________________

 

Record the class data here…        Red _____    Yellow ______         Blue ______