BIOLOGY – 1ST
SEMESTER
TYPE OF LAB - MICROSCOPY
INTRODUCTION TO MICROSCOPY
Lab Format: This lab is a remote lab activity Microscopy.
Relationship to Theory: In this lab you will learn the underlying principles that allow a
microscope to function and you will learn to operate a microscope.
Instructions for Instructors: This protocol is written under an open source CC BY license. You
may use the procedure as is or modify as necessary for your class. Be sure to let your students
know if they should complete optional exercises in this lab procedure as lab technicians will not
know if you want your students to complete optional exercise.
Remote Resources: Primary - Microscope, Secondary – introduction to Microscopy slide set.
Instructions for Students: Read the complete laboratory procedure before coming to lab. Under
the experimental sections, complete all pre-lab materials before logging on to the remote lab,
complete data collection sections during your on-line period, and answer questions in analysis
sections after your on-line period. Your instructor will let you know if you are required to complete
any optional exercises in this lab.
Contents
Learning Objectives ..................................................................................................... 2
Background Information ............................................................................................... 2
Equipment ................................................................................................................... 6
Preparing to Use the Remote Web-based Science Lab (RWSL) ................................. 6
Introduction to the Remote Equipment and Control Panel ........................................... 7
Experimental Procedure: ............................................................................................. 7
Pre-Lab Exercise 1: The Microscope ........................................................................... 7
Exercise 2: Operating a Compound microscope ......................................................... 8
Pre-Lab Exercise 3: Field of View and Depth of Field of a Compound Microscope ... 11
Exercise 3: Field of View and Depth of Field of a Compound Microscope ................. 11
Pre-Lab Exercise 4: Observation of Cells .................................................................. 12
Exercise 4: Observation of Cells ................................................................................ 12
Summary Questions: Introduction to Microscopy....................................................... 13
Appendix A - Introduction to the RWSL Microscope .................................................. 15
Appendix B – Loading Slides ..................................................................................... 16
Appendix C - Microscope Control .............................................................................. 17
Appendix D – Manipulating the Microscope Image .................................................... 18
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Appendix E – Capturing and Saving a Microscope Image ......................................... 20
Appendix F - Camera Controls .................................................................................. 22
LEARNING OBJECTIVES
After completing this laboratory experiment, you should be able to do the following things:
1.
2.
3.
4.
Identify the parts of a compound microscope and operate it effectively.
Understand the basic differences between Dissecting, Compound, and Electron microscopes.
Demonstrate competency with the focus and operation of a microscope.
Use a microscope to capture images and turn them into figures.
BACKGROUND INFORMATION
There are few things that have impacted the development of modern biology more than the
development of the microscope. Not only did the microscope open up a whole new world of
observation, which directly led to the discovery of microorganisms, cells, and some cellular
components. The microscope affected the thought process and conduct of biological research. As
Sachs wrote in History of Botany “… the use of the magnifying glass brought an advantage with it of
a different kind - it taught those who use them to see scientifically and exactly.”(Julius Sachs, 1890)
Additionally, 3 Nobel prizes have been awarded for the development of microscopes; the
ultramicroscope won the award for Chemistry in 1923, phase contrast microscopy won the award
for Physics in 1953, and electron and force microscopes won the award for physics in 1986 (Nobel
Organization). Since the microscope is such an important instrument not only for scientific
observation but for the development of the field of biology we will spend a little time on the history
and development of the instrument.
Humans have had at least a basic understanding of lenses for a long time. The existence of “burning
glasses” and “magnifying globs” (hollow glass spheres filled with water) can be traced back almost
2000 years if not more(Singer, 1914). However, it was not until the 1300s, around the time we begin
to see lenses used for eye glasses, that magnification started to be used by biologists(Singer, 1914).
These earliest devices were single lens microscopes and had limited magnification compared to
modern scopes. The pinnacle of this type of microscope was Leeuwenhoek’s discovery of “bacteria”
in 1673(Mazzarello, 1999).
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During the 1600s we begin to see the creation of the compound microscope, which is a microscope
with multiple lenses. The earliest of these microscopes were very similar, if not identical, to
telescopes, being simple tubes with two or
more lenses.
The first practically used
compound microscope was invented by the
Janssens in 1590(Singer, 1914).
The
compound microscope provided a much
higher degree of magnification than the
earliest scopes, however at cost in clarity due
to chromatic aberration and image distortion
at higher magnification. This area of
microscope development is most notably
remembered for Hook’s detailed analysis
published
in
his
Micrographia
in
1665(Mazzarello, 1999) this is the work that
coined the term “cells.”
The time period starting in the early 1800s to
present has been predominantly focused on eliminating distortions and aberrations so that
compound microscopes could be pushed to their absolute resolution limit(Singer, 1914). We will
return to a discussion of the resolution limit of a compound microscope later. The first optical
aberration we will discuss is chromatic aberration. This type of defect is caused because white light
is actually composed of many different colors of light. As an example consider a prism. On one side
white light goes in and on the other a rainbow comes out (Figure 1). This is because the degree of
refraction (the bending of the light) is dependent on the wavelengths of the light, the shorter
wavelengths are refracted less than the longer wavelengths. In this case the prism is a basic lens.
Therefore in a compound microscope that uses white light, the different wavelengths that compose
this light would focus at different points because they are each refracted a different amount. This
produces a color blur around objects (chromatic aberration). Over many years this problem was
fixed with the development of compound lenses, which are composed of multiple lenses. In this case
each of the sub-lenses has a different refractive index. This brings the focal point of the different
wavelengths of light back together(Ernst, 1900). These are called achromatic or apochromatic
lenses.
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The other type of distortion is curvilinear distortion which is the tendency to bend straight lines into
curves. An example that you are likely familiar with is the fisheye effects seen in wide angle
photography. The reason for the distortion is that light bends as it strikes a lens at a nonperpendicular angle. In order to get the high degree of magnification needed in early compound
microscopes, the single lenses that were used had very curved surfaces. This meant that the light at
the edge of the lens was bent much more than the light from the center of the lens. To correct this,
microscope builders again used compound lenses. In this case, the sub-lenses have different
shapes(Ernst, 1900). The different shapes will affect the bending of the light rays so that the light
rays are brought back perpendicular to each other. Since the shape of a lens changes its effect on
light, multiple lenses can be used to shape light in many different ways; Figure 2 shows the effect of
different lenses on light rays.
Over the last several decades the development of modern microscopes has begun to be focused on
the resolution limit. The resolution of a microscope is defined as the ability of the observer (person
or camera) to distinguish between two objects that are close together. In 1873 Ernst Abbe showed
that the theoretical resolution limit of a diffraction microscope was half the wavelength of the light
being used(Lipson, 2011). Therefore the resolution limit of an optical microscope is about 190 – 350
nm (the human eye responds to light with wavelengths of 390 – 700 nm). To get better resolution
than is possible with a compound microscope, scientist started using electron beams with scanning
and tunneling electron microscopes that have wavelengths 100,000 times smaller than visible
light(Scherzer, 1949). Recently, work using lasers and quantum mechanics has begun to develop
microscopes that “break” the resolution limit of optical microscopes, however, we will not discuss
these types of microscopes further in this protocol.
Since the resolving power of a microscope also indicates how much magnification you theoretically
have, the type of microscope you use depends on the type of observations you are conducting. We
will now briefly cover the other types of microscopes you should be familiar with. In addition to the
compound microscope, you should be familiar with the dissecting microscope and electron
microscopes.
Of the three types of microscope, the dissecting microscope has the lowest magnification. The
magnification range of a dissecting microscope is 10x to 40x. It provides a view of a fairly large sample
for the purposes of detailed examination and/or dissection. Typical subjects might be small animals,
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whole flowers, or organ parts. Dissecting microscopes are used to magnify specimens of sizes 10 μm
to 0.1 m. If a dissecting microscope has two ocular lenses on separate body tubes, it is a stereoscope
dissecting microscope. The dissecting microscope uses visible light which is brought in from the side
of the sample.
The compound light microscope has a magnification range that place it between the dissecting
microscope and the electron microscopes. The magnification range is 40x to 2000x. Typical subjects
might be cells, large cellular organelles, and bacteria. Compound microscopes are used to magnify
specimens of sizes 200 nm to 5 mm. The
compound microscope uses visible light
which is brought in from below the sample.
Since the light must shine through the
specimens, it must be small or thinly sliced to
obtain a good image. Since you will be using
this type microscope exclusively in this lab
we will also discuss the components and
parts of this microscope. In a modern
compound microscope there are three main
optical components (Figure 3). They are the
condenser (Figure 3a), objective (Figure 3b)
and the eye piece (Figure 3c). The condenser
focuses the light from the light source onto
the underside of the sample. The objective
contains a complex series of lenses that
correct chromatic aberration and image
distortion. The objective also gives part of the
microscope’s total magnification.
The
objectives are mounted into a wheel that
allows the user to select which objective they
wish to use. The length of the objective is
often related to its magnification with longer
objectives having higher magnifications. One
of the reasons that the objectives are different
lengths is so that the microscope will be parfocal at all magnifications, which means that the object that is in focus at 10X will also be in focus at
20X, 40X, 60X etc. The eye piece gives the final part of the magnification such that the total
magnification of a microscope is determined by multiplying the objective by the eye piece. As an
example if you’re using a 10X eyepiece and 20X objective your total magnification will be 10 x 20 =
200X.
The electron microscopes have the highest effective magnification of the microscopes we are
discussing. Electron microscopes come in two types The scanning electron microscope (SEM) and
the transmission electron microscope (TEM) each type employs electron bombardment to image
very small specimens. In the SEM the electrons pass through a three dimensional specimen and are
“read” using a detection device. A computer reconstructs the specimen image from the information
gathered by the scanning process. The magnification range of an SEM can reach 200,000X, and
provide great detail. With the TEM you can allows internal investigations the internal structure of
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prepared specimens. The TEM has a higher magnification range than the SEM, up to 50,000,000X.
Electron microscopes are used to image samples that range from 1 nm to 100 μm in size.
References:
Ernst, H.C. (1900). THE DEVELOPMENT OF THE MICROSCOPE. J. Boston Soc. Med. Sci. 4, 148–152.1.
Julius Sachs, I.B.B. (1890). History of Botany (1530-1860) (Clarendon press).
Lipson, A. (2011). Optical physics (Cambridge ; New York: Cambridge University Press).
Mazzarello, P. (1999). A unifying concept: the history of cell theory. Nat. Cell Biol. 1, E13–E15.
Organization, N.P. Microscopes: Time Line.
Scherzer, O. (1949). The Theoretical Resolution Limit of the Electron Microscope. J. Appl. Phys. 20, 20–29.
Singer, C. (1914). Notes on the Early History of Microscopy. Proc. R. Soc. Med. 7, 247–279.
EQUIPMENT
 Paper
 Pencil/pen
 Slides
o Letter ”e”, Whole Mount
o Colored Threads, Whole Mount
o Stage Micrometer
o Elodea Leaf, Whole Mount
o Hydra, Longitudinal Section
o Mixed Protozoa, Whole Mount
 Computer with Internet access (for the remote laboratory and for data analysis)
PREPARING TO USE THE REMOTE WEB-BASED SCIENCE LAB (RWSL)
Click on this link to access the Install guide for the RWSL: http://denverlabinfo.nanslo.org
Follow all the directions on this webpage to get your computer ready for connecting to the
remote lab.
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INTRODUCTION TO THE REMOTE EQUIPMENT AND CONTROL PANEL
Watch this short tutorial video to see how to use the RWSL control panel:
http://denverlabinfo.nanslo.org/video/microscope.html
For a more in-depth description of all the functions of the control panel:
https://www.youtube.com/watch?v=yW_HtlJONoI
There are appendices at the end of this document that you can refer to during your lab if
you need to remind yourself how to accomplish some of the tasks using the RWSL
control panel.
EXPERIMENTAL PROCEDURE:
Once you have logged on to the microscope you will perform the following Laboratory procedures:
PRE-LAB EXERCISE 1: THE MICROSCOPE
There are many types of microscopes available for biologist to use in this day and age. The
functionality of these microscopes is dependent on many factors. Which type of microscope you use
is dependent on the particular task you are trying to accomplish.
Pre-lab Questions:
1. One of the most common types of microscope in use today is the compound light microscope.
What property of this microscope’s construction gives it the name “compound microscope”?
2. Complete the following table of magnifications
Objective
10X Eye Piece
20X Eye Piece
Total Magnification
4X
10X
20X
40X
60X
Use the microscope types in the following list to answer the following questions. The names may be
used more than once.
Dissection Microscope
Compound Microscope
Scanning Electron microscope
Transmission Electron Microscope
3. The _________________ microscope has the highest magnification.
4. The _________________ microscope is used to examine a whole organ.
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5.
6.
7.
8.
The _________________ microscope has two or more sets of lenses.
The _________________ microscope uses light from the side to illuminate its sample.
The _________________ microscope can generate magnifications of 200,000X.
The _________________ microscope used to look at whole cells or large cellular organelles.
9. Label the parts on the following diagram of a compound microscope.
10. You are trying to build a light microscope with the best resolution possible. To do this your
light source is going to be a single wavelength of visible light. You have single wavelength
light-emitting diodes in Infrared, Red, Orange, Green, Blue, and Violet. Which one will you use
as your light source and why?
EXERCISE 2: OPERATING A COMPOUND MICROSCOPE
In this exercise you will learn how to operate a compound light microscope. When the exercise is
over you will understand how to position a sample, focus the microscope, and change the
magnification.
Data Collection:
1. Select the letter e slide from the microscope interface.
2. Place the letter e in the center of the field of view and Focus on it with the 10X objective.
First, we are going to examine the effects of magnification.
3. In the space below is a diagram of the slide as it is currently loaded on the microscope (Figure
4 a). In box (Figure 4 b) draw the letter e as you see it in the microscope.
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4. What are the differences between the observed letter e and the letter e mounted on the
microscope? (hint there are two differences)
5. Change to the 20X objective and capture an image of the letter e. Paste that image below.
6. Change to the 60X objective and capture an image of the letter e. Paste that image below.
Second, we are going to examine microscope movement.
7. Change back to 10X objective.
8. Now using the microscope controls, click on the top button which moves the stage in (towards
the body of the microscope) so that the e moves slightly. How does the e move with respect
to the direction the stage moves?
9. This time, press the left stage control button this will move the stage to the left (towards the
slide loader) so that the e moves slightly. How does the e move with respect to the direction
the stage moves?
Analysis:
10. Describe the difference in your observations of the black lines that make up the letter e using
the 20X and 60X objectives.
11. In the three diagrams below assume that each dashed line represents the distance the stage
will move with each click of the control buttons. Using the sequence of buttons draw where
the e will be after the button clicks.
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PRE-LAB EXERCISE 3: FIELD OF VIEW AND DEPTH OF FIELD OF A COMPOUND MICROSCOPE
When you are examining a sample under the microscope you need to know both how much of the
sample and what part of it you are viewing. We determine how much of the sample we are viewing
based on the amount visible to the ocular or the camera, this area is called the field of view. The field
of view is a plane parallel to the slide and can be easily quantified using a ruler. While you are viewing
a sample the part you see is the part of the sample that is in the focal point. There may be parts of
the sample either above or below the focal point that you cannot see. The distance that is in focal
point is called the depth of field. The depth of field is perpendicular to the field of view. However
depth of field is much harder to measure then field of view and we will only be dealing with depth of
field qualitatively in this lab. Both field of view and depth of field are related to and affected by the
magnification of the objectives. In this section you will explore the relationship between the objective
magnification and the field of view and depth of field.
Pre-lab Questions
1. How do you think the depth of field and field of view will change as magnification increases.
Scientist utilize a specific methodology when conducting experiments, this process is called the
scientific method. Vary simple in the scientific method you ask a question, make a prediction, test
that prediction, and then revise your prediction if necessary. Your answer to question one in this
exercise is a prediction and the reaming parts of this exercise will allow you to test and revise your
prediction. However, scientists craft their predictions in a specific form called a hypothesis. In order
for a hypothesis to a scientific hypothesis it must be both logically valid and testable. One way to
write a hypothesis is to use an If … Then … statement, an example of which would be If the field of
view decreases in diameter Then the depth of field will increase in size. Using this information
complete the next pre-lab question.
2. Rewrite you your answer to the previous question into an If … Then … hypothesis.
EXERCISE 3: FIELD OF VIEW AND DEPTH OF FIELD OF A COMPOUND MICROSCOPE
Data Collection:
First, we are going to examine the depth of field.
3. Select the Overlapping Threads slide from the microscope interface.
4. Examine the slide with the 10X objective and describe what you see.
5. How many of the threads can you get in focus using the 10X objective? Which of the threads
is on top, in the middle, and on the bottom? Capture and image and, using your word
processor, label the top middle and bottom thread. Paste it below
6. Change to the 20X objective, how many of the threads can you get in focus simultaneously?
7. Change to the 40X objective, how many of the threads can you get in focus simultaneously?
8. Change to the 60X objective, how many of the threads can you get in focus simultaneously?
Capture and image and paste it below.
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Second we will examine the field of view.
9. Select the Stage Micrometer slide from the microscope interface. A stage micrometer is a
microscope slide that has a small ruler drawn on it. In this case the ruler is 1mm in total
length with 100 divisions.
10. What is the length of one division on the stage micrometer?
11. Next you are going to measure the diameter of the field of view for each of the 5 objectives
the microscope has. Create a table to record the diameter of the field of view for each of the
objectives.
12. Select each objective in turn and measure the diameter across the middle of the visible space,
each group member should collect a set of data. Paste your table with all the groups data
below.
Analysis:
13. What happens to the depth of field as the magnification increases?
14. Take the table you created in question 11 and add two columns to it. In the first column
average the diameter measurements for all your groups’ data. In the second column convert
the mm lengths into µm lengths.
15. What happens to the field of view as the magnification increases?
16. Refer back to the If … Then … hypothesis you created in Pre-Lab Exercise 3, question 2. Was
your prediction correct? Explain why or why not.
17. Rewrite your hypothesis in light of the information you learned performing this experiment.
PRE-LAB EXERCISE 4: OBSERVATION OF CELLS
In the next exercise we will look at actual biological samples. The slides we will use are: Elodea Leaf,
Hydra, and Mixed Protozoa. We will use these samples to make observations using the microscope
and examine the differences between different types of cells.
Pre-lab Questions:
1. Do you expect to see any differences between the cells in the Elodea Leaf and the Hydra? What
differences do you expect to see?
2. Rewrite your answer to question 1 in the format of an If … Than … hypothesis.
3. Do you think the Protozoa will be more like plants or animals? Why?
4. Rewrite your answer to question 3 in the format of an If … Than … hypothesis
EXERCISE 4: OBSERVATION OF CELLS
Data Collection:
5. Select the Elodea Leaf from the microscope interface.
6. Center and focus the sample in the field of view. Change to the 40X or 60X objective. Capture
an image of the leaf cells. Paste the image below.
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In the previous exercise we determined the diameter of your field of view. We will use this
information to determine the size of the Elodea cells.
7. Change your objective to 40x and select an area on the slide so that the leaf cells completely
fill the field of view. Create a table to record your data.
8. Position the slide so that the bottom edge of a cell is at the bottom edge of your field of view.
Count how many cells are present in the cell column to reach the top edge of your field of
view.
9. Repeat step 8 three more times with different columns of cells. Paste your table below.
10. Select the Hydra slide from the microscope interface.
11. Center and focus the sample in the field of view. Change to the 40X or 60X objectives. Capture
an image of the Hydra cells. Paste the image below.
12. Select the Mixed Protozoa slide from the microscope interface.
13. Find some protozoa on the slide. Center and focus the sample in the field of view. Change to
the 40X or 60X objective. Capture an image of the protozoa. Paste the image below.
Analysis:
14. Using your word processor and the image you captured in step 6, label all the structures you
can identify in a leaf cell (examples: nucleus, cell membrane, cytoplasm, cell wall etc.). Paste
the image below.
15. Next we are going to determine the size of the leaf cells. Start by averaging the number of cells
it takes to fill your field of view from top to bottom. Insert that number below.
16. Next divide the diameter of your field of view by the average number of cells. This will give
you the size of your cell. Insert that number below (make sure to include your units).
17. Using your word processor and the image you captured in step 11, label all the structures you
can identify in a Hydra cell (examples: nucleus, cell membrane, cytoplasm, cell wall etc.).
Paste the image below.
18. Did you identify any differences between plant and animal cells? How do these differences
compare to your hypothesis?
19. Using your word processor and the image you captured in step 13 label all the structures you
can identify in a protozoa cell (examples, nucleus, cell membrane, cytoplasm, cell wall etc.).
Past the image below.
20. Based on your observation about the protozoa, are protozoa cells more like plants or animal
cells? Explain.
Until fairly recently all organisms were classified into one of the five kingdoms of life (Monera, Fungi,
Protista, Plantae, and Animalia). This classification system was based on physical appearance and in
some cases physiology. More recent studies using DNA sequence comparison has shown that all
living organisms belong to three Domains of life Bactria, Archaea, and Eukaryota. In this system the
it was realized that Plants, animals, and Protista all belong to the Eukaryota domain.
21. In light of the idea that Plants, animals, and Protista are all separate groups of the Eukaryota
domain does your observations about the protistas make more sense? Why?
SUMMARY QUESTIONS: INTRODUCTION TO MICROSCOPY
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1. You are looking at a sample at 10X when you increase the magnification to 40X the sample is
no longer in the field of view. Why might this happen and how would you correct it?
2. If the compound microscope is par-focal explain why sometimes when you go from the 10X
or 20X object to the 60X object is the sample not in focus.
3. If an electron microscope has the highest resolution and magnification why do we still use
dissecting and compound microscopes?
4. Why does an electron microscope have a higher magnification and resolution than a
compound light microscope?
5. Given the following slide draw how the sample would appear while looking through the
microscope.
6. It is important to understand how light moves through a microscope. Complete the diagrams
in figure 7 to show the light path, in figure 7a draw the mirror that is needed to direct light to
the ocular instead of the camera. In figure 7b draw the path that the light takes to reach the
ocular. In figure 7c draw the path the light takes to reach the camera.
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APPENDIX A - INTRODUCTION TO THE RWSL MICROSCOPE
The RWSL microscope is a high-quality digital microscope located in the remote lab
facility. You will be controlling it using a control panel that is designed to give you
complete control over every function of the microscope, just as if you were sitting in front
of it.
You must call into a voice conference to communicate with your lab partners and with
the Lab Technicians. This is very important because only one person can be in control
of the equipment at any one time, so you will need to coordinate and collaborate with
your lab partners.
You take control of the equipment by right-clicking anywhere on the screen and
selecting Request Control. You release control by right-clicking too.
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APPENDIX B – LOADING SLIDES
Clicking on the Slide Loader tab at the top
of the screen will display the controls for
the Slide Loader robot. There can be up to
four cassettes available on the Slide
Loader, and each cassette can hold up to
50 slides. There will be a drop-down list
for each cassette that is available. In the
above example, only cassette #1 is
available on the Slide Loader. You can
click on it to select a specific slide to be
loaded, as in the image below:
Once you select the slide you want to load
on the microscope, click the Load button to
the right of the drop-down list. You will see
a message telling you that the slide is
loading. You can also watch this
happening using the picture-in-picture
(PIP) camera (see Appendix F - Camera
Controls).
Notice that when a slide is actually on the
microscope (or when it is being loaded or
unloaded), the cassette controls will be
grayed out so you cannot load a second slide
until the first is removed.
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Once the slide is on the microscope, it will be
listed in the “Current Slide on Stage” box,
and the only thing that the Slide Loader robot
can do is return it to the cassette when you
click the “Return Slide to Cassette” button.
To move the slide around while it is on the
microscope stage, you must return to the
Microscope tab to see those controls.
APPENDIX C - MICROSCOPE CONTROL
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The microscope stage controls are boxed in red in the above image. The allow you to
move the microscope stage (which holds the specimen slide) left, right, forward or
backward. You can also focus by moving the stage up and down.
You can change the objective, which gives you increased or decreased magnification, by
clicking the buttons under Objective Selection.
The Condenser control controls whether or not the Condenser lens is in the light beam.
You want to have the condenser OUT for the 4x objective, but IN for all the others.
APPENDIX D – MANIPULATING THE MICROSCOPE IMAGE
You can manipulate the microscope image by using the controls in the red boxed area
above. The White Balance should be used only if the image appears to be brown or
gray and you think you might need to adjust it (although it won’t hurt anything to click
this button).
The Normal, Negative, etc, control buttons in this area are used to display the image
slightly differently in order to highlight certain features. Here is some information from
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the Nikon website (http://www.microscopyu.com/articles/digitalimaging/digitalsight/correctingimages.html)
about these settings and when they might be used:
Normal: In this mode, the image is displayed in the natural color scheme that is observed in the microscope eyepieces (Figure 3).
For the majority of images captured with the Digital Sight system, the normal color output is the most effective mode for accurate
and effective reproduction of all specimen details.
Negative: The Negative effect displays a brightness- and color-inverted form of the image, where red, green, and blue values are
converted into their complementary colors (Figure 4). The technique is useful with specimens for which color inversion can be of
benefit in exposing subtle details, or in quantitative analysis of specimens.
Blue Black: This mode represents the black portions of the Negative image in blue, and is often useful to reveal details in
specimens having a high degree of contrast. As a special effect, the Blue Black mode can be beneficial as a presentation tool.
Black & White: This mode displays a grayscale form of the image (Figure 6). It can be effectively used for monochromatic images
such as those acquired with differential interference contrast or phase contrast techniques. In many cases, digital images destined
for publication in scientific journals must first be converted into black & white renditions of those captured in full color. The B &
W filter can often aid the microscopist in preparing images for publication or oral presentation.
Sepia: This effect is essentially a monochrome image version displayed in sepia (brownish) tones instead of grayscale (Figure
7). The Sepia mode is more likely to be utilized in general photographic applications than in microscopy, although the effect may
enhance the visibility of specimen detail in certain instances.
Auto Exposure is normally turned on, but you can turn it off if you want to play around
with the brightness of the light source and not have the microscope camera automatically
adjust, though it’s usually best to leave it turned on.
If you turn off the Auto Exposure, then some new controls appear that let you turn the
LED off or on, and also adjust the intensity of the light source. The intensity of the light
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source can be increased or decreased manually with the dial that now appears next to
the Objective control.
APPENDIX E – CAPTURING AND SAVING A MICROSCOPE IMAGE
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2
You can capture a high-resolution image of what is currently in the field of view of the
objective by clicking the Capture Image button, which will turn bright green while it is
capturing the image. When the Capture Image light turns off, the image has been
successfully captured. After the image is captured, click View Captured Image to see the
high-resolution image (below).
After opening this image, right click on it and select “Copy”. Then paste it into a document
so you can use it later in your lab report. This is illustrated below.
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After right-clicking and selecting Copy, just open a document and right-click and select
Paste. You can either paste it directly into your lab report document or into another one
for safe-keeping until you use it later.
You can use drawing tools in your editor to annotate this image so you can show your
instructor that you knew what you were supposed to be looking for!
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APPENDIX F - CAMERA CONTROLS
Clicking the Picture-in-Picture button will open a window that shows the view from a
camera placed directly in front of the microscope. The arrow buttons allow you to swivel
the camera around so you can see whatever you want to look at in the lab. The
Camera Preset Position buttons are programmed to show you particular portions of the
apparatus. If you hover the mouse over them, a box will pop up that lists what each
position will show you (see below).
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