Week 5
9/24/13 - 9/26/13
in week 5 of class we performed an acid-fast stain, an endospore stain, and determined if our bacteria was motile or not. We first started with the acid-fast staining. An acid-fast stain is used to distinguish the two groups of bacteria based on the lipid content and their cell walls: acid-fast and non-acid-fast. We first made a bacterial smear on a glass slide and then fixed it over a Bunsen burner. We then placed the slide over a beaker of boiling water. We took a piece of bibulous paper and set it on the slide before dropping carbolfuchsin over the length of the slide. We continued to drop carbolfuchsin on the slide for 3 minutes.
We then removed the bibulous paper and retrieved the slide using forceps. We rinsed the slide using water to remove excess stain and then a decolorizing solution, adding drop by drop until the color stopped running. We then immediately rinsed the slide to remove the decolorizing agent. We then covered the slide with methylene blue for 2 minutes.
Then we rinsed the slide with water to remove the excess coloring. After blotting the slide with pieces of bibulous paper we examined the smear under the microscope
We then removed the bibulous paper and retrieved the slide using forceps. We rinsed the slide using water to remove excess stain and then a decolorizing solution, adding drop by drop until the color stopped running. We then immediately rinsed the slide to remove the decolorizing agent. We then covered the slide with methylene blue for 2 minutes.
Then we rinsed the slide with water to remove the excess coloring. After blotting the slide with pieces of bibulous paper we examined the smear under the microscope
upon examination we determined that are bacteria was a non acid-fast due to the methylene blue stain
We then were given a tube with motility test medium. Using aseptic technique, we took a sample of our bacterial culture with an inoculating needle and then stabbed it into the test medium. We then placed it in the incubator at 37' C until the next lab period.
The following lab, we worked on preparing an endospore stain. This stain is used to determine if a particular bacteria has endospores. We again fixed a smear of bacteria on a glass slide. We then placed the smear over a beaker of boiling water. Taking a piece of bibulous paper and placing it over the smear, we dropped malachite green over the smear. We let the stain set for 6 minutes while continuing to add stain as it evaporated.
We then discarded the paper and retrieved the slide using forceps. We rinsed the slide with water to remove the excess dye. We then covered the smear with safranin for 70 seconds. We immediately rinsed the slide with water to remove the excess safranin and after blotting the slide with pieces of paper we examine the smear under the microscope.
We then discarded the paper and retrieved the slide using forceps. We rinsed the slide with water to remove the excess dye. We then covered the smear with safranin for 70 seconds. We immediately rinsed the slide with water to remove the excess safranin and after blotting the slide with pieces of paper we examine the smear under the microscope.
upon examination we determine that are bacteria did not contain endospores
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