Title: preparation of films for staining Objective: to produce a thin smear of bacteria adhering to a clean microscope slide as preparatory to staining. Procedure: A. From broth cultures 1. Inoculating loop was sterilized using Bunsen burner and let to be cool before use it to obtain bacterial suspension from the tube. 2. The bacterial then placed in the center of the clean microscope slide. It also speared to produce thin films. 3. The films are set to air dry before wafting the slide gently on the Bunsen flame to fix and kill pathogenic bacteria. 4.
The slide was place on a rack then the stain were apply. B. From agar cultures. 1. A drop of water was place in the center of the glass slide. 2. With sterilized inoculating loop to obtain a minute of bacterial culture from the agar cultures 3. It then mix with the drop of water and then proceeds as broth cultures. SIMPLE STAINING TECHNIQUES Material: 1. 24 hours broth cultures of a. E. coli b. Staph. aureus 2. 24 hours nutrient agar slants of a. Bacillus subtilis b. Pseudomonas aeruginosa 3. Slides 4. Inoculating loop 5. Dye solution a. Crystal violet b.
Methylene blue c. Carbol fuchsin 6. Test tubes Procedure: 1. By using inoculating loop a minute of bacterial was transferred onto the glass slide. 2. Then a film of slide was prepared. 3. The prepared film then flooded with crystal violet, methylene blue,and carbol fuchsin. All of the stain it let to react about 30 seconds. 4. The excess stained was wash with slow running water. The excess water then blot before dry with Bunsen flame. 5. The slide then examine without the cover slip under the objective lens (x100) 6. The observation then recorded. Question
In preparing a slide from a colony growing on an agar medium, why is it necessary to place only a minute portion of the colony on the slide? Answer To obtain a good slide from a colony growing on an agar medium it is necessary to place only a minute portion of bacterial colony. Because a thick film will reduce the light penetration on the slide that result error or not accurate images on the structure that we observed. On the other hand, thick film will make the structure we observed in clumps image or double or three layers, this will affect our observation.
Therefore, it is really important to use only a minute portion of bacterial colony on the slide. Discussion: From the experiment, we know the simple stain is used to differentiate the shape of the bacteria, the arrangement, and also the size of the bacteria. The contrast between the one bacterium with another bacterium is poorly observed. Conclusion: In this experiment we are able to produce a thin smear of bacteria adhering to a clean microscope slide as preparatory to staining.