Lab -2

Microscope with unprepared slides

2.1 Objective:
In this lab, we will explore the fundamentals of using a microscope. We'll begin by
familiarizing ourselves with each part of the microscope and understanding what it does. Next,
we'll learn how to calculate magnification and explain what happens when we adjust the field
of view. We'll then practice locating objects using both low and high magnification settings.
Finally, we'll demonstrate how to prepare a slide and identify what we see on it, starting with
an unprepared slide.
2.2 Introduction:
It refers to an optical instrument that uses a lens or an arrangement of lenses to magnify an
object. Also, they help to view different organisms.
The compound microscope, which consists of at least two lenses, was invented in 1590 by
Dutch spectacle-makers Zacharias and Hans Jansen. Some of the earliest microscopes were also
made by a Dutchman named Antoine Van Leeuwenhoek.
Leeuwenhoek’s microscopes consisted of a small glass ball set inside a metal frame. He became
known for using his microscopes to observe freshwater, single-celled microorganisms that he
called “animalcules.” Its magnification is 1000X
An electron microscope is a microscope that uses a beam of electrons as a source of
illumination. They use electron optics that are analogous to the glass lenses of an optical light
microscope to control the electron beam, for instance focusing them to produce magnified
images or electron diffraction patterns. It can magnify things up to millions of times. Its main
types are TEM and SEM.
2.3 Literature review:
2.3.1 Parts of microscope:
Head: The head is a cylindrical metallic tube that holds the eyepiece lens at one end and
connects to the nose piece at other end. It is also called a body tube or eyepiece tube
Arm: This is the part connecting the base to the head and the eyepiece tube to the base of the
microscope.
Base: The base is the lowermost part of the microscope that supports the entire microscope
structure. It provides stability for the microscope.
Eyepiece: The eyepiece (ocular Lens) is closest to the viewer’s eye. They are located at the top
of the microscope. This part is used to look at the specimen. These lenses come in different
magnification powers from 5X to 30X, but the most common ocular lenses are of 10X or 15X
magnification. They magnify the image for the second time.
Eyepiece tube: It’s the eyepiece holder. It carries the eyepiece just above the objective lens.
Nose piece: A nose piece is a movable circular structure that houses all the objective lenses. It
is also called the revolving turret. It is connected to the body tube and lies just above the stage
Objective lenses: The objective lens is the lens that is closest to the specimen. They are fitted
on the nosepiece. A standard microscope has 3 to 4 objective lenses of different magnifying
powers: 4X, 10X, 40X, and 100X. The objective lenses first receive the light transmitted from
the specimen and magnify the image for the first time.
The Adjustment knobs: Adjustment Knobs are the control knobs used to focus the
microscope on the specimen.
 Fine Adjustment Knob: Fine Adjustment Knob is used for fine adjustment. It is a
smaller knob and is used to move the stage up or down very slowly.
 Coarse Adjustment Knob: Coarse Adjustment Knob is used for focusing the image
under low power magnification. It is a larger knob and is used to move the stage up or
down very rapidly. The stage is raised or lowered rapidly with the help of a coarse
adjustment knob.
Aperture: This is a hole in the microscope stage through which the transmitted light from the
source reaches the stage.
Condenser: These are lenses that are used to collect and focus light from the illuminator into
the specimen. They are found under the stage next to the diaphragm of the microscope. They
play a major role in ensuring clear, sharp images are produced with a high magnification of
400X and above.
2.3.2 Magnification and resolving power:
2.4 Methodology:
The first step to preparing your microscope slide is to gather all required instruments. Now that
you’ve gathered your materials, you can move on to mounting the slide.
It’s important to note that there are a few different types of slide mounts
Preparing dry mount slide:
A dry mount slide is the easiest to prepare. The only steps needed are to place a specimen on
the slide, and then place a coverslip over it if needed. This is easier than preparing a wet mount
slide since a liquid is not required and there is no risk of air bubbles that can make it difficult to
clearly view the specimen.
Preparing wet mount slide:
A wet mount slide requires liquid, such as water or oil. A benefit to adding a liquid to a slide is
to help magnify the specimen. The first step for preparing a wet mount slide is to place the
specimen on the slide. Then, add a drop of liquid to the slide using a dropper pipette. Next, set
one edge of the coverslip on the slide next to the specimen and slowly lower it until it is lying
flat. This will help prevent air bubbles, allowing for a clearer view of the specimen. It might be
helpful to use tweezers to hold the coverslip as you lower it. Stains may also be applied to help
view certain cell structures.
Preparing smear mount:
 Prepare a small sample of your specimen.
 Transfer the sample onto a slide.
 From a broth culture, use a sterile pipette to transfer 1-2 drops onto your slide. Spread
the drop into an even smear spread thinly across the length of your slide.
 From a solid media culture, use a sterile pipette to transfer a drop of sterile water or
saline solution to the middle of a slide. Using a sterile inoculation loop, isolate a small
sample of the culture and mix evenly into the drop of sterile water or saline solution.
 Spread the drop into an even smear spread thinly across the length of your slide.
 Allow the smear to dry completely before undergoing fixation to adhere the smear to
the slide via heat fixation or methanol fixation.
  Using a stain helps improve a specimen’s contrast, making features more distinguishable
under a microscope. Staining a sample is especially useful for classification as it will
highlight a specimen’s shape, as well as distinct features such as a nucleus in eukaryotes.
 Adjust the microscope and make it stable
 Use the fine adjustment knob to bring the specimen into sharper focus. Be gentle when
adjusting the focus, as turning the knob too quickly or too hard can damage the
mechanism.
 Examine your specimen. Select the appropriate objective lens to view your samples.
They allow you to magnify the sample at different levels, typically 4x, 10x, 40x, and
100x.
2.5 Results:

Guard
cells

Stomata

Figure 2.1: shows leaf epidermis, showing guard cells, stomata

Figure 2.2: shows leaf epidermis

2.6 Conclusion:
In this lab, we're going to learn how to use a microscope. We'll start by getting to know
each part of the microscope and what it does. Then, we'll figure out how to make things look
bigger and how changing what we see affects our observations. We'll also practice finding tiny
things using the microscope and learn how to prepare slides and identify what's on them.
2.7 Questions:
1. Take two different samples for study.
Sample 1 Leaf epidermis Sample 2 Onion epidermis

Figure 2.3: leaf epidermis Figure 2.4: onion epidermis

Leaf Epidermis:
1. Shape: Leaf epidermis cells appear irregular in shape with uneven edges.
2. Structures: Some leaf epidermis cells may have stomata (tiny pores) and guard cells that
regulate gas exchange and transpiration.
3. Patterns: Depending on the type of leaf, you may observe various patterns such as veins
and trachoma’s (hair-like structures) on the surface.

Onion Epidermis:
1. Shape: Onion epidermis cells are typically rectangular or square-shaped with clear
boundaries.
2. Layers: Onion epidermis often shows multiple layers of cells stacked on top of each
other.
3. Appearance: The cells of onion epidermis are densely packed and usually lack any
distinct structures like stomata or veins
2. Use the microscope to observe the prepared slides at different magnifications (e.g.,
4x, 10x, 40x, and 100 x). Note any changes in clarity and level of detail. On which
objective lens your image is best. Also calculate magnification power.
Observing prepared slides through the microscope at various magnifications (like 4x, 10x,
40x, 100x) reveals changes in clarity and detail. The image is clearest and most detailed under
 the 40x objective lens. Magnification is calculated by multiplying the magnification of the
objective lens by that of the eyepiece.
3. Prepare two sets of slides for the same specimen—one stained and one unstained.
Compare and contrast the observations between the two sets.
Comparing stained and unstained slides of the same specimen shows distinct differences
in visibility and contrast. The stained slides reveal clearer outlines and internal structures due to
the dye highlighting specific features. Unstained slides, on the other hand, offer a more natural
view but may lack clarity in certain details. This comparison underscores the importance of
staining techniques in enhancing visualization and aiding in the identification of microscopic
structures.
4. Draw the diagram of specimen you observed under the microscope. Also label it.

5. Conduct a group discussion on the observations made during the lab, sharing
insights and comparing findings with classmates.
In our group discussion, we shared our observations from the lab, exchanging insights and
comparing findings with each other. We found that different specimens showed varying levels
of clarity and detail under different magnifications. We also discussed the differences between
stained and unstained slides, noting how staining techniques enhanced visibility. Additionally,
we shared tips and techniques we found helpful during slide preparation and specimen
identification. Overall, the discussion was valuable in deepening our understanding of
microscopy and improving our skills for future experiments.
2.8 References
 https://www.toppr.com/guides/biology/microbiology/microscope-types-uses-parts/
 https://www.microscopeworld.com/p-3658-types-of-microscopes.aspx
 https://www.microscopeworld.com/t-parts.aspx
 https://sciencing.com/magnification-microscope-5049708.html