Gram Stain Experiment

Gram Stain: The important part of this experiment is being able to determine a bacterium based on its cell wall structure. It also helps indentify if the unknown organism is a Gram positive or Gram negative. This is the initial step that must be taken before any other lab procedure may continue on to ensure the purity is present, the arrangement of the cells, and the shape the cell has. The staining of the cell starts off with using the primary stain, Crystal Violet which is a purple color, to begin with. Next, Gram Iodine is added on, which is our mordant and this causes a complexion with the primary stain, results in formation of insoluble complexes. Then, a counter stain is applied on called Safranin (pinkish/red color). In Gram positive
The agar contains Triphenyl tetrazoliumchloric (TTC) which gives off a color indicator for motility. If the organism moves from the stab indention, then the inoculation shows it was motile and grew red in the agar. However, if the organism did not move from the stab indention, then it will appear just a thin precipitate of red, which indicates a non-motile growth. Compare the motility with the positive control bacteria, Proteus vulgaris, and negative control bacteria, Staphylococcus aureus, to gauge the movement of the organism. Refer to Brown, Microbiological Applications
Mycobacterium). These organisms contain mycolic acid in their cell walls rather than peptidoglycan. The use of Acid alcohol produces the stain since it does not decolorize an acid-fast organism. The acidic dye, Carbol Fuchsin, is the primary stain to use in order to retrain the acid-fast organism’s cell wall. Next, the basic counterstain, Methylene blue, is added on to stain the rest of the organism with the expectation of the acid-fast bacterium. This will help create a color contrast of the acid-fast organism. The stain procedure is outlined in Brown, Microbiological Applications