INTRODUCTION
- To contrast unstained and stained bacteria.
- To note differences in the study of living and dead microorganisms
- To stress again precautionary measures to be observed in microbiological laboratory.
METHODOLOGY:
A. Demonstration of Brownian movement
The oscillating or quivering motion exhibited by all very minute particles suspended in the water fluids is known as the Brownian movement.
- Place a drop of carmine red in glass slide. Slowly lay a cover slip over it to avoid bubbles forming in the slide.
- Examine under the microscope and observe.
B. Hang-drop preparation and microscope examination
1. Spread a little glycerin around the edge of the cover slip.
2. With a platinum loop, transfer a loopful of broth culture or fluid to be examined as a culture growing on a solid culture medium or material. Place a drop of sterile seawater (SSW) in the center of the cover slip and mix with a loopful of inoculum.
3. Place the depression slide over the cover slip in such a way that the center of the depression slides lies over the drop.
4. Note that the glycerin seals the cover slip to the slide, holds it in place and prevents evaporation.
5. Quickly invert the slide. The drop to be examined hangs from the undersurface of the concavity at any point.
6. Be sure that the platinum loop is flame-sterilized every time it is used.
7. Examine the hang-drop with 16mm objective to focus it them swing to 4mm objective or high dry. Reduce the amount of light passing through the penetration by partly closing the iris diaphragm of the substage condenser of the microscope. A properly made preparation shows the bacterial cell standing out distinctly against a dimly lighted background. Brownian movement and the flowing of organisms and particles in current s on the fluid must not be mistaken. True motility depends on the possession of flagella. If so equipped they can propel themselves with progressive directional locomotion.
8. Compare your result with what you observe during the demonstration of the Brownian movement.
9. Discard the hang-drop preparation into suitable container of disinfectant after exercise.
Guide Questions for Discussion
1. Give the usefulness of the hang drop.
2. Give the precautionary measures of handling and disposing hang-drop preparations.
3. Describe the Brownian movement. Indicate how it may be distinguished from true motility?
4. What means of locomotion do bacteria possess?
5. Describe the motility observed in hang-drop preparation you have made of bacteria. Which bacteria are motile?
6. Why must sterile saline solution is used in making hang-drop from solid media?
REFERENCES
Laboratory Exercise No. 7 Direct Examination of Unstained Bacteria
