LAB 2: MEASUREMENT AND COUNTING
OF CELLS USING MICROSCOPE
2.1
OCULAR MICROMETER
INTRODUCTION
To measure and compare
the size of prokaryotic and eukaryotic microorganisms, we need to used ocular
micrometer. Ocular micrometer is inserted into the one of the microscope
eyepiece to measure microorganisms. The micrometer , which serve as a scale or rule, is a flat
circle of glass upon which are etched equally spaced division. This is not calibrated,
and may be used at several magnifications. When placed in the eyepiece,the line
superimposed certain distance markers on the microscope field. The actual
distance superimposed may be calibrated using a stage micrometer on which
parallel lines exactly 10um apart etched.
By determining how many units of the ocular micrometer superimpose a
known distance on a stage micrometer, you can calculate the exact distance each
ocular division measure on the microscopic field. When changed the objectives
we must recalibrate the system. After calibration of the ocular micrometer, the
stage micrometer is placed with a slide containing microorganisms. The
dimension of the cells can be determined.
OBJECTIVE
1.
To
measure cells using microscope
2. To
determine how many units of ocular micrometer superimpose a known distance on
the stage
micrometer.
3.
To
calculate the exact distance each ocular division measure on the microscopic
field.
RESULT
Yeast
400x magnification
1000x magnification
Lactobacillus
400x magnification
1000x magnification
DISCUSSION
We using ocular micrometer
to measure and compare the size between yeast and Lactobacillus. Yeast is a
type of eukaryotic and Lactobacillus is prokaryotic. Size of eukaryotic
typically much bigger than prokaryotic. We use 400x and 1000x magnification to
observe the microorganisms after the calibration to get the size of
microorganisms.
From the experiment we
get:
|
Magnification
|
400x
|
1000x
|
||
|
Calibration
|
Stage
micrometer:
|
Ocular
division :
|
Stage
micrometer:
|
Ocular
division:
|
|
0.03 mm
|
38
|
0.09 mm
|
96
|
|
|
0.00079
mm
|
1
|
0.0094
mm
|
1
|
|
|
0.79 um
|
1
|
0.94 um
|
1
|
|
|
Yeast:
400x magnification : 4
division x 0.79 um = 3.16 um
1000x magnification : 7 division x 0.94 um =6.58 um
Lactobacillus:
400x magnification : 2 division x 0.79 =1.58 um
1000x magnification : 5 division x 0.94 =4.70 um
| ||||
From this result we can
conclude that Lactobacillus is much smaller than yeast. Lactobacillus are the
types of prokaryote which have a nuclear region which contain DNA, and
ribosomes, the site of protein synthesis, as their main features. They have a
cell membrane but do not have an internal membrane system.
CONCLUSION
As a
conclusion, we can measure and compare the size of prokaryotic and eukaryotic
microorganisms using ocular micrometer. Example of prokaryote is Lactobacillus
and for eukaryote is yeast. Calibration is used to determine the units of
ocular micrometer superimpose a known distance on the stage micrometer. The
size of prokaryote is much smaller than the size of eukaryote.
REFERANCE
1 .
Pearson
International Edition Biology, 8th Edition, Campbell, Reece
2 .
International
Student Edition Biochemistry, Campbell, Farrell
2.2
NEUBAUER CHAMBER
INTRODUCTION
Neubauer
chamber is used to count microbes. The Neubauer is a heavy glass slide with two
counting areas separated by a H-shaped trough. A special cover slip is placed
over the counting areas and sits a precise distance above them.
OBJECTIVE
1.
To
count cells using microscope.
RESULT
Yeast, 400x magnification
DISCUSSION
In this experiment to avoid contamination
of microorganisms to the culture or us getting infection from it we must do
aseptic technique. We used flame
sterilization to the media bottle containing yeast and used 70% ethanol to
clean the Neubauer and cover slip as well. Sterile Pasteur pipette is used to
transfer the a drop of dilute yeast culture to the space between the cover slip
and the counting chamber. To prepare the counting chamber the mirror-like polished surface is carefully cleaned with lens paper. The coverslip is also cleaned .Coverslip for counting chambers are specially made and are thicker than those for conventional microscopy, since the must be heavy enough to overcome the surface tension of a drop liquid.The coverslip is placed over the counting surface prior tp put on the cell suspension.The suspension is introduced into one of the V-shaped wells with the pasteur. The area under the coverslip fills by capillary action.Enough liquid should be introduced so that the mirrored surface is ust covered. The charged counting cahmber is placed on the microscope stage and the counting grid is brought into focus at low power until 400x magnification.
From the experiment, we can see
that:
56 ÷ 10 = 5.6
= 6
Volume = 0.05 x 0.05 x 0.01 mm
= 2.5 x 10⁻⁵ mm
= 2.5 x 10⁻⁷ ml
6 cells in 16 squares
1 square = 6 ÷16
= 0.375 cells in 2.5 x 10⁻⁷ ml
1ml = 0.375 ÷ 2.5 x 10⁻⁷
= 1500000 cells/ml
CONCLUSION
Neubauer chamber is the best
technique to count microbes. Aseptic technique is used to protect user from
infection, prevent contamination and prevent toxicity to the culture. For a
reasonably accurate count, we should have more than 30 cells per area.
REFERANCE
1.
Pearson
International Edition Biology, 8th Edition, Campbell, Reece
No comments:
Post a Comment