Applying Staining Techniques To View And Identify

Bacteria Essay, Research Paper Abstract The main objective of this lab was to identify different bacteria by simple, negative, and gram staining. To view each bacteria cell, the bacteria was transferred aseptically to a slide, and they were then viewed by using oil immersion, by a light microscope. From this lab, it was determined that E. coli and B. megaterium are gram negative and B. subtilis and S.

Bacteria Essay, Research Paper

Abstract

The main objective of this lab was to identify different bacteria by simple, negative, and gram staining. To view each bacteria cell, the bacteria was transferred aseptically to a slide, and they were then viewed by using oil immersion, by a light microscope. From this lab, it was determined that E. coli and B. megaterium are gram negative and B. subtilis and S. Marcesans are gram positive.

Introduction

The purpose of this lab was to view the different characteristics of bacteria by applying various staining techniques. It is important to know the make up if a certain bacteria so an antibiotic may be engineered to destroy the bacteria. From the gram stain, it was possible to determine which bacteria was gram positive or gram negative. This is important because gram-negative bacteria are generally more toxic (due to the lipopolysaccaride) are resistant to antibiotics than the gram-positive bacteria.

Methods

The materials used for this lab were:

1. A light microscope

2. Four glass slides

3. And inoculating loop

4. A Bunsen Burner

5. Bacteria (E.coli, S. megaterium, B. subtilis, and S. marcesens)

6. Alcohol

7. Staining bowl

8. Methylene blue, oil, nigrosin, crystal violet, iodine, safranin

9. Distilled water

Three different staining procedures were then used for all four types of bacteria. The directions for each staining process can be found on pages 18-19 of the lab manual. For simple stain, bacteria was removed with a sterile inoculating loop and placed on a glass slide. Methylene blue was then applied until the slide was covered. Distilled water was then poured on the slide until the methylene blue was removed. The slide was allowed to dry. Next, oil was placed on the slide so that the oil objective lens on the light microscope was employed. The bacterium was viewed and a sketch was made. For the negative stain, a loopful of bacteria was placed on a slide. A drop of Nigrosin solution was placed next to the bacteria, and the blood smearing technique learned in the first lab was applied to cover the slide with nigrosin solution. A sketch was then made. Gram staining was more complicated. First, two separate types of bacteria were used. One type was placed on one slide, and the other bacteria were placed on the opposite side. Both bacteria were also placed in the middle of the slide so they could be viewed together. Crystal violet, iodine and safranin were all placed on the slide and washed off with distilled water. Once dried, oil was applied to the slide, and it was ready to be viewed. Again a sketch was made.

Results

The sketches were drawn on a separate piece of paper and then revised. A table is

provided.

E.coli B. subtilis S. marcesans S. megaturiun

Size Small, rod-shaped Small, rod-shaped Round, very small Round, small clumps

Elevation Convex Flat Convex Convex

Surface Shiny Rough Red, smooth Smooth

Color White, shiny Pale yellow Reddish-orange Light purple

Edge Wrinkled Round Round Round

Discussion

The basic purpose of this lab exercise was to view the different characteristics of certain types of bacteria. Using the oil immersion lens accomplished this task. The importance of the lens is that it can get so close to the slide (it is basically touching it) and gives the experimenter a great view of the desired object. The different staining techniques were very useful. The simple stain just stains the cells, so the basic shape of the cell is viewed. With the negative stain, the background is stained, so the elevation, color, and surface are more easily observed. The gram stain, dyed the bacteria either red or purple by using the iodine, methylene blue, crystal violet, and safranin, helped determine the difference between the gram negative or gram-positive bacteria. If the gram stain is negative it will have a lipopolysaccharide cell wall, if positive, it is a peptidoglycan cell wall. Once this is known, scientists can engineer antibiotics that disrupt the bacteria’s process of protein synthesis. Some different examples of this are erythromycin and tetracycline.

References

Appendix- Gene Transfer in the Environment.

Cambell, Neil. Biology. 4th Edition. Benjamin/Cummings Pub. Co. 1996

Abstract

The main objective of this lab was to identify different bacteria by simple, negative, and gram staining. To view each bacteria cell, the bacteria was transferred aseptically to a slide, and they were then viewed by using oil immersion, by a light microscope. From this lab, it was determined that E. coli and B. megaterium are gram negative and B. subtilis and S. Marcesans are gram positive.

Introduction

The purpose of this lab was to view the different characteristics of bacteria by applying various staining techniques. It is important to know the make up if a certain bacteria so an antibiotic may be engineered to destroy the bacteria. From the gram stain, it was possible to determine which bacteria was gram positive or gram negative. This is important because gram-negative bacteria are generally more toxic (due to the lipopolysaccaride) are resistant to antibiotics than the gram-positive bacteria.

Methods

The materials used for this lab were:

1. A light microscope

2. Four glass slides

3. And inoculating loop

4. A Bunsen Burner

5. Bacteria (E.coli, S. megaterium, B. subtilis, and S. marcesens)

6. Alcohol

7. Staining bowl

8. Methylene blue, oil, nigrosin, crystal violet, iodine, safranin

9. Distilled water

Three different staining procedures were then used for all four types of bacteria. The directions for each staining process can be found on pages 18-19 of the lab manual. For simple stain, bacteria was removed with a sterile inoculating loop and placed on a glass slide. Methylene blue was then applied until the slide was covered. Distilled water was then poured on the slide until the methylene blue was removed. The slide was allowed to dry. Next, oil was placed on the slide so that the oil objective lens on the light microscope was employed. The bacterium was viewed and a sketch was made. For the negative stain, a loopful of bacteria was placed on a slide. A drop of Nigrosin solution was placed next to the bacteria, and the blood smearing technique learned in the first lab was applied to cover the slide with nigrosin solution. A sketch was then made. Gram staining was more complicated. First, two separate types of bacteria were used. One type was placed on one slide, and the other bacteria were placed on the opposite side. Both bacteria were also placed in the middle of the slide so they could be viewed together. Crystal violet, iodine and safranin were all placed on the slide and washed off with distilled water. Once dried, oil was applied to the slide, and it was ready to be viewed. Again a sketch was made.

Results

The sketches were drawn on a separate piece of paper and then revised. A table is

provided.

E.coli B. subtilis S. marcesans S. megaturiun

Size Small, rod-shaped Small, rod-shaped Round, very small Round, small clumps

Elevation Convex Flat Convex Convex

Surface Shiny Rough Red, smooth Smooth

Color White, shiny Pale yellow Reddish-orange Light purple

Edge Wrinkled Round Round Round

Discussion

The basic purpose of this lab exercise was to view the different characteristics of certain types of bacteria. Using the oil immersion lens accomplished this task. The importance of the lens is that it can get so close to the slide (it is basically touching it) and gives the experimenter a great view of the desired object. The different staining techniques were very useful. The simple stain just stains the cells, so the basic shape of the cell is viewed. With the negative stain, the background is stained, so the elevation, color, and surface are more easily observed. The gram stain, dyed the bacteria either red or purple by using the iodine, methylene blue, crystal violet, and safranin, helped determine the difference between the gram negative or gram-positive bacteria. If the gram stain is negative it will have a lipopolysaccharide cell wall, if positive, it is a peptidoglycan cell wall. Once this is known, scientists can engineer antibiotics that disrupt the bacteria’s process of protein synthesis. Some different examples of this are erythromycin and tetracycline.

Bibliography

References

Appendix- Gene Transfer in the Environment.

Cambell, Neil. Biology. 4th Edition. Benjamin/Cummings Pub. Co. 1996

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