Gram Stain Test Lab Report

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I. Purpose:

The purpose of this lab was to learn how to properly mount a bacteria for examination under a microscope. Also learned in this lab were methods on how to inoculate an agar or broth medium, and using aseptic techniques. Lastly, how to use stains to visualize bacterial cells, or distinguish different types of cells from one another through the usage of dye techniques such as the Gram-stain test was learned.

II. Prepping/Mounting/Simple Staining Bacterial Cells Procedure:

1. Obtain 3 glass slides that are free of any foreign matter.
2. Place one drop of water on the slide.
3. Touch an inoculating loop to the petri dish of the desired bacteria and rub the loop onto the slide with the drop of water.
4. Let the slide air dry.
5. Do
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Rinse again with deionized water for 5 seconds.
7. Dry the slide and observe at the 100x objective, write down observations such as color and shape.

Potassium Test Procedure:

1. Take a drop of KOH.
2. Get a blob of bacteria using an inoculating loop.
3. Mix the blob of bacteria using the inoculating loop, on the slide for 30 seconds.
4. Observe what happens to the bacteria, and check the results against what is known to happen to certain kinds of bacteria in the presence of KOH, Gram positive will turn watery and will have a purple color, while Gram negative will be mucosal and turn pink.

Prepping Test Tube Bacterial Specimens for Spectrophotometer Use/Using a Spectrophotometer Procedure:

1. Blank the spectrophotometer.
2. Get a small cuvette.
3. Use a pipette to add the mixture you want to use in the spectrophotometer, and add the mixture to the cuvette.
4. Vortex the cuvette until it is completely mixed and uniform.
5. Place cuvette into the spectrophotometer.
6. Make observations about what the spectrophotometer says about the absorbance of the bacterial broth in the cuvette and write down the results. The reading should give a number value for absorbance.

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None of the cultures that were inoculated with Kocuria rosea were pink as they should be, and this could indicated contamination either because of improper inoculation techniques, such as the loop being too hot when introduced to the Kocuria rosea, leading to them being killed off, or the petri plate could have been left open too long to foreign materials, leading to bacterial growth that was not intended. Also, the inoculating loop could have not been sterilized correctly with the Bunsen burner. Making sure the loop is not too hot when doing the inoculation, leaving the petri plate as closed as possible when working with it, and also making sure the loop is kept in the flame for a longer amount of time, are all things that could be done to improve the outcome of this experiment in the

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