Lab 4271

LAB 1: NUR DIANA BT ABDUL JALIL(114120) LAB 1 : PRINCIPLES AND USE OF MICROSCOPE

1.1 Setting up and using the microscope

Introduction

A microscope (from the Greek: μικρός, mikrós, "small" and σκοπεῖν, skopeîn, "to look" or "see") is an instrument used to see objects that are too small for the naked eye. The science of investigating small objects using such an instrument is called microscopy.
Microscopic means invisible to the eye unless aided by a microscope. Microscopes magnify microscopic specimens and make them appear larger than they truly are. This allows us to accurately study and describe cells and microscopic organisms.

The Part of The Microscope

· eyepiece-where you look through to see the image of your specimen.

· body tube-the long tube that holds the eyepiece and connects it to the objectives.

· nosepiece-the rotating part of the microscope at the bottom of the body tube; it holds the objectives.

· objective lenses-(low, medium, high, oil immersion) the microscope may have 2, 3 or more objectives attached to the nosepiece; they vary in length (the shortest is the lowest power or magnification; the longest is the highest power or magnification).

· arm-part of the microscope that you carry the microscope with.

· coarse adjustment knob-large, round knob on the side of the microscope used for focusing the specimen; it may move either the stage or the upper part of the microscope.

· fine adjustment knob-small, round knob on the side of the microscope used to fine-tune the focus of your specimen after using the coarse adjustment knob.

· stage-large, flat area under the objectives; it has a hole in it (see aperture) that allows light through; the specimen/slide is placed on the stage for viewing.

· stage clips-shiny, clips on top of the stage which hold the slide in place.

· aperture-the hole in the stage that allows light through for better viewing of the specimen.

· diaphraghm-controls the amount of light going through the aperture.

· light or mirror-source of light usually found near the base of the microscope; the light source makes the specimen easier to see.

Results

· Stained Cells

Streptococcus sp. - 40x

Streptococcus sp. - 1000x

Streptococcus sp.- 4000x

· Wet Mount

Saccharomyces arevisiae sp. (yeast) – 1000x

Lactobacillus fermentum sp. - 1000x

Discussion

Bacteria is too tiny to be discovered by low-power or high-power dry objectives. Instead they are stained and observed with the oil immersion objectives that being used only at very large magnifications that require high resolving power. Objectives with high power magnification have short focal lengths, facilitating the use of oil. The oil is applied to the specimen (conventional microscope), and the stage is raised, immersing the objective in oil.

Another method in used in the examination of unstained bacteria with the use of the microscope is the wet mount method. Wet mounts can detect the characteristics of most types of known bacteria. The method classifies bacteria by type as being gram positive, gram negative or gram variable. The classification of the bacteria in question is determined by the color that is seen under the microscope after the chemical stain has been added to the mount. The color that is exhibited by specific bacteria is directly related to the bacterial cell wall's ability to retain the color during examination.

Saccharomyces Cerevisiae (Yeast)

Saccharomyces cerevisiae, which in Latin means “sugar fungus,” has been utilized by humans for thousands of years. It is believed that it was first discovered on the skins of grapes. S. cerevisiae is a budding or brewing yeast, and has been put to use since antiquity to make dough rise and to provide ethanol in alcoholic beverages.

Its deeply detailed cellular structure makes S. cerevisiae, also known as "brewers yeast," one of the most highly researched model organisms in the study of biology. It exists in single-cell form, or in pseudomyceliac form. Cellular reproduction occurs by budding. The ability of S. cerevisiae to ferment specific sugars is a major factor that differentiates it from other yeasts.

S. cerevisiae exists and grows in the haploid and diploid cellular forms. The haploid life cycle consists of mitosis, growth, and ultimately death, the latter more rapid under extremely stressful conditions. Diploid cells also undergo mitosis as well as growth, but in the same stressful circumstances can experience sporulation. Subsequent to sporulation, the cells undergo meiosis and produce a number of haploid spores. These haploid spores progress to mate.

Lactobacillus Fermentum

Lactobacillus fermentum is a Gram-positive species of bacterium in the genus Lactobacillus. It is associated with active dental caries lesions. It is also commonly found in fermenting animal and plant material. It has been found in sourdough. A few strains are considered probiotic or "friendly" bacteria in animals and at least one strain has been applied to treat urogenital infections in women. Some strains of lactobacilli formerly classified as Lactobacillus fermentum (such as RC-14) have since been reclassified as Lactobacillus reuteri. Commercialized strains of L. fermentum used as probiotics include PCCand ME-3. Lactobacillus fermentum belongs to the genus Lactobacillus. Species in this genus are used for a wide variety of applications. These applications include food and feed fermentation.

Conclusion

Morphology of the microorganism can be observed by microscope by oil immersion objectives which requirevery high resolving power for large magnifications. Besides, wet mount method also can be used in the examination of unstained bacteria.

Reference

· http://biology.clc.uc.edu/fankhauser/labs/microscope/Oil_Immersion.htm

· http://www.ruf.rice.edu/~bioslabs/methods/microscopy/oilimm.html

· http://biologicalmicroscopy.com/examination-of-unstained-bacteria.html

· http://microbewiki.kenyon.edu/index.php/Lactobacillus

· http://www.google.com.my/search?ix=sea&sourceid=chrome&ie=UTF-8&q=morphology+of+lactobacillus+fermentum

LAB 1 : PRINCIPLES AND USE OF MICROSCOPE by Nurul Samihah binti Mohd Jamil (111408)

Introduction:

Anthony van Leeuwenhoek

Robert Hooke

The microscope is perhaps the most basic tool used by biologists. Light microscopy was first applied to biological materials in the seventeenth century by such researchers as Robert Hooke and Anthony van Leeuwenhoek. The impact of this invention on the course of human endeavor is easily underestimated. Within a short span of time, that which was invisible to the human eye became visible. An entire world of living organisms previously unknown was now available for study. The microscope remains a pivotal tool in learning biology to this day, with most laboratories being equipped with one or more research instruments.

The part of the microscope;

Body Tube-The body tube holds the objective lenses and the ocular lens at the proper distance

Nose Piece-The Nose Piece holds the objective lenses and can be turned to increase the magnification

Objective Lenses-The Objective Lenses increase magnification (usually from 10x to 40x)

Stage Clips-These 2 clips hold the slide/specimen in place on the stage

Diaphragm-The Diaphragm controls the amount of light on the slide/specimen

Light Source-Projects light upwards through the diaphragm, the specimen and the lenses ; Some have lights, others have mirrors where you must move the mirror to reflect light

Ocular Lens/Eyepiece-Magnifies the specimen image

Arm-Used to support the microscope when carried. Holds the body tube, nose piece and objective lenses

Stage-Supports the slide/specimen

Coarse Adjustment Knob-Moves the stage up and down (quickly) for focusing your image

Fine Adjustment Knob-This knob moves the stage SLIGHTLY to sharpen the image

Base-Supports the microscope

Magnification-To determine your magnification…you just multiply the ocular lens by the objective lens ; Ocular 10x, Objective 40x:10 x 40 = 400x ; So the object is 400 times “larger” ; Ocular lenses usually magnifies by 10x

Using a Microscope-Start on the lowest magnification ; Don’t use the coarse adjustment knob on high magnification…you’ll break the slide!!! ; Place slide on stage and lock clips ; Adjust light source (if it’s a mirror…don’t stand in front of it!) ; Use fine adjustment to focus

The Bright-Field Microscope-produces a dark image against a brighter background ; has several objective lenses- parfocal microscopes remain in focus when objectives are changed ; total magnification- product of the magnifications of the ocular lens and the objective lens

Objectives:

Learn to use a simple bright-field microscope correctly
Learn the parts of a compound light microscope and their functions.
Learn how to calculate the magnification of a compound light microscope.
Learn how to make a wet mount slide.
Understand how the orientation and movement of the specimen’s image changes when viewed though a compound light microscope.
Learn the proper use of the low and high power objective lenses.
Learn the proper use of the coarse and fine adjustments for focusing.

Results:

Stained Cell

Species: Streptococcus sp. ;
Magnification : 40x

Species: Streptococcus sp. ;
Magnification : 100x

Species: Streptococcus sp. ;
Magnification : 400x

Wet mount

Species: Saccharomyces cerevisiae ;
Magnification : 1000x

Species: Lactobacillus fermentum ;
Magnification : 1000x

Discussion:

Morphology,

in biology, the study of the size, shape, and structure of animals, plants, and microorganisms and of the relationships of the parts comprising them. The term refers to the general aspects of biological form and arrangement of the parts of a plant or an animal. The term anatomy also refers to the study of biological structure but usually suggests study of the details of either gross or microscopic structure. In practice, however, the two terms are used almost synonymously.

Stained Cell

Streptococci are a diverse genus, infecting a barrage of different animals, including humans, with diseases ranging from strep throat to necrotizing fasciitis.

They have come to public attention recently as antibiotic-resistant strains have started appearing and causing epidemics. In an effort to battle the evolution of these clever pathogens, researchers have sequenced the genomes of 9 different strains in 4 different species of Streptococcus.

Streptococci are nonmotile, Gram-positive, nonsporeforming bacteria, that live in pairs or chains of varying length. They are characteristically round or ovoid in shape. Most Streptococci are facultative anaerobes, although some are obligate anaerobes.

They usually require a complex culture medium in order to grow. Many streptococci imitate aspects of their host in order to escape detection. The capsule of Streptococcus pyogenes is chemically similar to that of it's host's connective tissue, and therefore, is nonantigenic, and it's cytoplasmic membrane has antigens similar to human cardiac skeletal and smooth muscle.

Wet mount

Saccharomyces cerivisiae is a species of yeast. It has developed as a model organism because it scores favorably on a number of these criteria.

As a single celled organism S. cerevisiae is small with a short generation time (doubling time 1.25–2 hours at 30 °C or 86 °F) and can be easily cultured. These are all positive characteristics in that they allow for the swift production and maintenance of multiple specimen lines at low cost.
S. cerevisiae can be transformed allowing for either the addition of new genes or deletion through homologous recombination. Furthermore, the ability to grow S. cerevisiae as a haploid simplifies the creation of gene knockouts strains.
As a eukaryote, S. cerevisiae shares the complex internal cell structure of plants and animals without the high percentage of non-coding DNA that can confound research in higher eukaryotes.
S. cerevisiae research is a strong economic driver, at least initially, as a result of its established use in industry.

Lactobacillus fermentum is a Gram-positive species of bacterium in the genus Lactobacillus. It is associated with active dental caries lesions.

L. fermentum belongs to 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 L. fermentum have natural resistances to certain antibiotics and chemotherapeutics. They are considered potential vectors of antibiotic resistance genes from the environment to humans or animals to humans.
Lactobacillus fermentum can also be a normal inhabitant of the human intestinal tract and some strains have been associated with cholesterol metabolism.

Conclusion:

We learnt the correct way to view sample of microorganisms by using simple bright-field microscope ( used different magnification 40x, 100x, 400x, 1000x to observe the microorganisms). We able to identify that how the orientation and movement of the specimen’s image changes when viewed though a compound light microscope.From the image obtain through microscopy, we are able to illustrate the diversity of cells and microorganisms and the description of morphology of the cells can be done.We examined a stained cell Streptococcus sp. , Saccharomyces cerevisiae, and Lactobacillus fermentum is the first experience in using light microscope.We used a lot of material and method to conduct this experiment and wet mount such as microscope, slides, cover slips, lens paper, and oil.

References:

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

Species ; Saccharomyces cerevisiae

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]

- A 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://en.wikipedia.org/wiki/Microscope

http://biology.touchspin.com/Bacteria_Coccus.php

http://www.biologyreference.com/La-Ma/Light-Microscopy.html#b

http://microscopyu.com/

http://www.molbiolcell.org/content/13/3/847.full.pdf

http://www.metamicrobe.com/lactobacteria/

http://tempedanbudu.blogspot.com

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