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Gram Stain Lab Report

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Materials Based off of the flow chart and the methods took to identify the two unknowns the tests I conducted were the Gram Stain Test, Endospore stain test, Urease test, Lactase test, and lastly the MRVP Methyl Red test. The first test conducted was the Gram Stain test. This test was used to determine if the two bacteria were either Gram Negative or Gram Positive. There are four main steps in the gram stain process that all affect the outcome and overall stain and accuracy of the test. The Primary stain is the first step in the gram stain. This process includes staining the cells, Negative and Positive cells, with Crystal Violet. At this point there is no difference in color of the stain, they are all purple. Next the Mordant step, you …show more content…

After figuring out that one bacteria is Gram Positive, the next step is to figure out if that bacteria has endoscopes or not. Only gram positive cells have the possibility of endospores, this also narrowing down the possible identities of the unknown bacteria. The endospore stain through a series of processes makes it so that only the endospores, if any, are stained green while the rest of the cells are stained pink. Allowing you to determine if there are any endospores and if so the shape and region of the spores. The staining process occurs through a series of four important steps. The Primary stain, Heat, Rinse, and Counter stain. The Primary stain, stains the cells and not the spores with a Malachite Green. Then, in the heat step, you heat the cells and spores over boiling water which allows the spores to expand and allow the Malachite to enter the spore. Now both cells are stained. The third step, rinse with water, allows the Malachite Green to rinse out of the cells but not the spores. It does not rinse from the spores because once the heated spores cool, they tighten up again and trap the stain inside. Lastly, adding Safranin allows the now clear cells to stain pink while the endospore stain stay green. Thus giving you the conclusion that there are round spores in the Gram Positive

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The unknown bacteria was then tested on multiple selective and differential media. Growth was present on the MacConkey Agar and the colonies were the same color as the plate, which told me my bacteria was gram negative and did not ferment lactose. There was no growth on the Mannitol Salt Agar, and this told me the unknown was not salt tolerant and did not

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The slide was then stained and left to steam with malachite green. It was continuously followed up by applications of the stain so it may remain moist for 10 minutes. The slide was then rinsed and safranin was again used as a counterstain. Using oil immersion objective lens of the microscope, unknown #76 had only reddish-pink cells without any signs of spore formation. Thus the given unknown is a non-spore former.

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