STRUCTURE: LABORATORY

Course Name & BWD 11730 GENERAL BIOLOGY

Code
Department Food Technology

Title Microscopy

Group Members 1.Nur Alya Farhanah Binti Kamalul Arifin (AW230082)

2.Nur Qurrotul Ain Binti Rosli (CW230017)
3.Nurul Zahidah Binti Nor Azmi (AW230037)
4.Muhammad Hafizul Bin Jamil (CW230055)
Section Section 1 / Group 1

Lecturer Name Muhammad Faiz Bin Razali

Submission Date 01 November 2023

INTRODUCTION

A microscope is an essential tool to see microorganisms that are too small to be seen by the
naked eye. There are many small objects or details of objects which cannot be seen by the
unaided human eye. The Microscopes are used to observe the shape of bacteria and host cells
in various stained and unstained preparations. A compound microscope is composed of two
elements : a primary magnifying lens and a secondary lens system. These experiments use 4x
lens, 10x lens, 40x lens and 100x lens and the total magnification of an image calculated by
multiplying the magnification of the ocular by the magnification of the objective. Study and
development use of microscopes throughout the ages and understanding of cells and their
structure and function has vastly improved. In cell biology, research and development
depended highly on the experimental methods to observe the behaviour and morphology of
cells. However, cells are too small to be seen with naked eyes. Therefore, Zaccharias Jansen
took the honour to invent the first compound microscope around 1600 (Tortora, Funke, &
Case, 2016). Following that, discovery of cells by Robert Hooke in 1665 was also aided by a
simple light microscope.

Furthermore, there are several microscopy techniques routinely used to study the cells.
Electron microscopes have higher magnification, higher resolution and more detail than light
microscopes. Transmission electron microscopy is similar to the observation of stained cells
with the bright field light microscope. Most cells are so tiny that they cannot be seen with the
naked eye because of that scientists use microscopes to study cells.
Figure 1.0 The microscope's parts and its functions.
 Table 1.0 microscope part’s and functions.

Microscope’s part Functions

Ocular lens where one view enlarges the object.

Body tube connect the eyepiece to the objective lenses.

Revolving nosepiece hold the objective lense ; rotate to change objective

lenses.

Objective lenses produce most of the magnifications.

Stage support the slide and contains an opening to allow light to

pass through the specimen into the objective.

Diaphragm rotating dial that controls the passage of light through the
stage.

Light source illuminates the specimen by shining bright light through

it.

Base support the entire microscope.

Stage clips hold the clips in place on the stage.

Arm support the lenses, mirror and body tube.

Fine adjustment knob used to make small focus adjustments to sharpen the
focus especially when using a high power objective lens.

Coarse adjustment knob moves the stage up and down quickly to find the
specimens when using low or medium power objective
lenses.

Power switch turn the light on or off

METHODOLOGY
1.1 The Bright Field Microscope

1. A prepared slide with the letter "e" on it was obtained. The slide was placed on the stage
and fixed with the slide holder.

2. The condenser focusing knob was rotated to move the condenser to its highest position.
The microscope was focused on any portion of the slide, then the condenser aperture was
closed down, and the condenser was moved until a sharply focused view was obtained..

3. The microscope was turned on by rotating the dimmer switch, and the light intensity was
adjusted to a comfortable level.

4. Looking down into the microscope, the eyepiece was adjusted according to our ocular
distance and diopter. The eye tubes were moved back or forth until one uniform field of view
was seen.

5. The microscope was focused at 10x magnification and then only went up to the desired
magnification.

6. With the slide firmly in place, the coarse focus control was rotated until the slide was as
close as possible to the 10X objective. The stage manipulators were moved until a portion of
the slide was directly under the objective and carefully focused on the object to be viewed.
After adjusting the focus at 10X, the object was centered to be viewed and the nosepiece was
rotated to the next higher magnification.

7. The fine focus was manipulated to get the sharpest image. During the use of the
microscope, one hand remained in fine focus as constant readjustment would be called for.
The other hand was used to manipulate the movement of the stage.

8. The slide was returned to the 10X objective and the slide was moved around until the letter
"e" in the view. The orientation of the letter "e" on the slide and in the field of view was
observed. The observations were recorded.

9. To use the 40X objective, the object was centered to view (the 40X will have a smaller
field of view) and the nosepiece was rotated to bring the 40X objective into position. The
changes in the orientation of the letter "e" was observed. The observations were recorded.

10. The image of the letter was drawn as observed under different magnifications.
1.2 Use Of Oil Immersion

1. The prepared slides were placed on the microscope stage. The microscope was focused on
an appropriate field containing bacteria using the 10X objective. The bacteria was centered in
the field of view by manipulating the lateral movement knobs.

2. The nosepiece was rotated to the 40X objective and refocused with the fine focus. The
object was centered again to be examined.

3. The nosepiece was rotated so that an intermediate position between the 40x and the 100x
objectives was obtained.

4. A small drop of immersion oil was placed on the center of the viewing area of the slide.

5. The nosepiece continued to rotate so that the 100X objective was rotated into the oil.

6. The shape of the bacteria was determined and drawn.

1.3 Measurements: Ocular and Stage Micrometers

1. A stage micrometer was placed on the stage of the microscope, and the lowest
magnification (4X) was used, and focused on the stage micrometer grid.

2. The ocular micrometer was rotated by turning the appropriate eyepiece. The stage was
moved until the superimpose the lines of the ocular micrometer over those stage micrometers.
With the two micrometer lines coinciding at one end of the field, the space of each
micrometer to a point at which the micrometer lines coincide again was counted.

3. The ocular divisions that are equivalent to one stage division where each division of the
stage micrometer measures 10 micrometers were counted. The number of micrometers in
each space of the ocular scale was calculated.

4. The process was repeated for 10X, 40X, and 100X. The calculations have been recorded.

5. Using the calculated values from the ocular micrometer, the dimensions of the letter "e" on
the microscope slide were determined, and the dimensions were added to the drawing in
Exercise 1.1. The letter "e" was measured directly with a millimeter ruler (without a
microscope) and compared with the calculated value you obtained through the microscope.
RESULTS :

1.1: The bright Field Microscope

Figure 1.1 : Letter “e” prepared slide under 10x objective lens

Figure 1.1.2 : Letter “e” prepared slide under 40x objective lens
1.2: Use of Oil Immersion (100x)

Figure 1.2: S. Aureus prepared slide under 100x objective lens

1.3: Measurement: Ocular and Stage Micrometers

Objective lens Wide Length Dimension

10x 100 120 12000

40x 100 70 7000

Stage Micrometer
1 division = 10µm (0.01 mm). Therefore
100 division x 10µm =1000µm

10x objective lens

100 ocular division = 1000µm
1 ocular division = 1000µm
100
= 10µm
Dimension = 10µm X 12000 = 120000µm

40x objective lens

Stage Micrometer
100 stage division x 10µm =1000µm
25 ocular division = 1000µm
1 ocular division = 1000µm
25
= 40µm
Dimension= 40µm X 7000 = 280000µm

Letter “e” measurement using a milimeter ruler = 0.1 mm

DISCUSSION :
Based on the result, the letter "e'' seen through the 10x objective lens appears larger and
inverted from its original position. Under the 40x objective lens, the letter "e" is larger than
10x, and only a straight line can be seen. The S.Aureus prepared slide can be seen by using
immersion oil under a 100x objective lens. The shape of S.Aureus is spherical. It appears as a
cluster of round cells. S.Aureus is classified as Gram-positive bacteria because it retains the
crystal violet stain and appears purple. The letter “e” value obtained using a millimeter ruler
is 0.1 mm. The calculated values obtained with a 10x objective lens are 120000µm, which is
less than the value obtained with a millimeter ruler, 0.1 mm, and the values obtained with a
40x objective lens. The calculated value obtained with a 40x objective lens is 28000µm,
which is less than the value obtained with a millimeter ruler. The field of view decreases as
magnification power increases.

The correct way to handle a microscope is to hold the microscope at the arm of the
microscope and put the other hand at the base of the microscope. Do not touch the lens with
bare hands. When examining images under a microscope, proper microscope handling is
critical. When using the microscope, make sure the slide clips hold the slide in place. All
prepared slides were examined at the lowest magnification of 4x, then 10x, 40x, and 100x.
For the 4x and 10x objective lenses, the coarse focus knob was used. For the 40x and 100x
objective lenses, the fine focus knob was used. Immersion oil is used under a 100x objective
lens to enhance resolution.

CONCLUSION:

A compound light microscope is a type of microscope that uses light to illuminate and
magnify specimens. It consists of several parts which are stage, light source, body tube, iris
diaphragm, condenser lenses, eyepiece, objective lenses, revolving nosepiece (turret), coarse
focus knob, fine focus knob, base and arm. Furthermore, the right steps to use a compound
light microscope is prepare the microscope, adjust the illumination, select the objective lens,
place the slide, center the specimen, start with coarse focusing, fine-tune the focus, adjust the
lighting, use the stage to controls the slide, observe and analyze, record observations, switch
off the microscope once finished observing and clean up the lenses.
REFERENCES
- Lab Manual BWD 11730 General Biology , UTHM
- https://www.scribd.com/document/492546434/BIO411-LAB-REPORT-1
- https://studylib.net/doc/9624327/microscope-lab-report
- https://sciencing.com/precautions-using-microscope-7379695.html