LAB 1 : PRINCIPLES AND USE OF MICROSCOPE by NOR SHAQIRA BT AZLAN (111391)


INTRODUCTION
A light microscope (LM) is an instrument that uses visible light and magnifying lenses to examine small objects not visible to the naked eye, or in finer detail than the naked eye allows.

The part of microscope.












Eyepiece Lens:  the lens at the top that you look through.  They are usually 10X or 15X power. 

Tube:  Connects the eyepiece to the objective lenses

Arm:  Supports the tube and connects it to the base

Base:  The bottom of the microscope, used for support

Illuminator:  A steady light source (110 volts) used in place of a mirror.  If your microscope has a mirror, it is used to reflect light from an external light source up through the bottom of the stage.

Stage:  The flat platform where you place your slides.  Stage clips hold the slides in place.  If your microscope has a mechanical stage, you will be able to move the slide around by turning two knobs.  One moves it left and right, the other moves it up and down.

Revolving Nosepiece or Turret:  This is the part that holds two or more objective lenses and can be rotated to easily change power.

Objective Lenses:  Usually you will find 3 or 4 objective lenses on a microscope.  They almost always consist of 4X, 10X, 40X and 100X powers.  When coupled with a 10X (most common) eyepiece lens, we get total magnifications of 40X (4X times 10X), 100X , 400X and 1000X.  To have good resolution at 1000X, you will need a relatively sophisticated microscope with an Abbe condenser.  The shortest lens is the lowest power, the longest one is the lens with the greatest power.  Lenses are color coded and if built to DIN standards are interchangeable between microscopes.  The high power objective lenses are retractable (i.e. 40XR).  This means that if they hit a slide, the end of the lens will push in (spring loaded) thereby protecting the lens and the slide.  All quality microscopes have achromatic, parcentered, parfocal lenses.

Rack Stop:  This is an adjustment that determines how close the objective lens can get to the slide.  It is set at the factory and keeps students from cranking the high power objective lens down into the slide and breaking things.  You would only need to adjust this if you were using very thin slides and you weren't able to focus on the specimen at high power. (Tip: If you are using thin slides and can't focus, rather than adjust the rack stop, place a clear glass slide under the original slide to raise it a bit higher)

Condenser Lens:  The purpose of the condenser lens is to focus the light onto the specimen.  Condenser lenses are most useful at the highest powers (400X and above).  Microscopes with in stage condenser lenses render a sharper image than those with no lens (at 400X).  If your microscope has a maximum power of 400X, you will get the maximum benefit by using a condenser lenses rated at 0.65 NA or greater.  0.65 NA condenser lenses may be mounted in the stage and work quite well.  A big advantage to a stage mounted lens is that there is one less focusing item to deal with.  If you go to 1000X then you should have a focusable condenser lens with an N.A. of 1.25 or greater.  Most 1000X microscopes use 1.25 Abbe condenser lens systems.  The Abbe condenser lens can be moved up and down.  It is set very close to the slide at 1000X and moved further away at the lower powers.  

Diaphragm or Iris:  Many microscopes have a rotating disk under the stage.  This diaphragm has different sized holes and is used to vary the intensity and size of the cone of light that is projected upward into the slide.  There is no set rule regarding which setting to use for a particular power.   Rather, the setting is a function of the transparency of the specimen, the degree of contrast you desire and the particular objective lens in use.
Magnification.
·         40x magnification
·         100x magnification
·         400x magnification
·         1000x magnification



OBJECTIVE
·         Learn to use a simple bright-field microscope correctly.
·         To provide an experience in the use of microscope.
·         To illustrate the diversity of cells and microorganisms.



RESULTS

Species ; Streptococcus sp
Magnification ; 40x

Species ; Streptococcus sp.
Magnification ; 100x

Species ; Streptococcus sp.
Magnification ; 400x


Magnification ; 1000x



Species ;   Lactobacillus fermentum 
Magnification ; 1000x




DISCUSSION
1. Stained cell
Cocci (plural of coccus) bacteria are found in circular smear. Cocci are too small to see below 10x, 100x and 400x. Although 1000x immersion oil magnification is done, individual cocci are still barely visible.
There are few types of arrangement of cocci shape ;
1.      Cocci [sing. Coccus]
- bacterial shape.
- Generally spherical though with some variation from this theme (i.e., elongation or flattening on one side).

2.      Diplococci
- Cocci that remain in pairs after they divide.

3.      Streptococci
- Cocci that fail to separate after they divide, but instead remain in chains of cells.

4.      Tetrad
- Cocci that fail to separate after they divide, but instead remain in groups of four forming squares

5.      Sarcinae 
 - Cocci that fail to separate after they divide, but instead remain in groups of eight forming cubes.

6.      Staphylococci
- Cocci that fail to separate after they divide, but instead remain in amorphous sheets or clumps.

2. Wet mount
i. Saccharomyces is a genus in the kingdom of fungi that includes many species of yeast.  Saccharomyces cerevisiae is used in making wine,bread and beer. Colonies of Saccharomyces grow rapidly and mature in three days.
They are flat, smooth, moist, glistening or dull, and cream to tannish cream in color. The inability to use nitrate and ability to ferment various carbohydrates.
A coccus-shaped bacterium is usually spherical, although some appear oval, elongated, or flattened on one side. Most cocci are approximately 0.5 - 1.0 micrometer (µm) in diameter and may be seen, based on their planes of division and tendency to remain attached after replication. .  Saccharomyces cerevisiae are commonly reproduce asexually by budding. Unlike bacteria which are prokaryote, Saccharomyces cerevisiae are eukaryotic.

ii. Lactobacillus fermentum is a Gram-positive species of bacterium in the genus Lactobacillus. Species in this genus are used for a wide variety of applications. These applications include food and feed fermentation. It has been found that some strains for Lactobacillus fermentum have natural resistances to certain antibiotics and chemotherapeutics.
 They are usually straight, although they can form spiral or coccobacillary forms under certain conditions. They are often found in pairs or chains of varying length.They can grow at temp. 45°C. Lactobacillus fermented belongs to arginine and ribose fermenting, a Betabacteria that are important in spoilage microorganism of beer. 

CONCLUSION
In conclusion, from this first lab, we learnt the basic technique and procedure to use the light microscope correctly. We also are now able to identify the specific function of each part of microscope. Examined a stained cell (Streptococcus sp.), Saccharomyces cerevisiae, and Lactobacillus fermentum is the first trial or experience in using light microscope. Different magnifications are used to obtain clearer image of the stained cell and the wet mount. From the image obtain through microscopy, we are able to illustrate the diversity of cells and microorganisms. Besides, the description of morphology of the cells can be done .


REFERENCES

http://mansfield.osu.edu/~sabedon/biol2010.htm#illustration_cocci


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