540 Q u E STI o NS 1 Predict the efect of the following -mistakes made when perf

Question

540 Q u E STI o NS 1 Predict the efect of the following -mistakes made when performing a Gram stain. Consider each a. Failure to add the iodine. Failure to the decoloriter apply Failure to apply the safranin. d. Reversal of crystal violet and safranin stains. 2 Both crystal violet and safranin are basic stains and may be used to do simple stains on Gram-positive and am-negative cells. This being the case,explain how they stain different cell types in the Gram stain. 3 If you saw large, eukaryotic cells in the preparation made from your gumline, they were most likely your oun epithelial cells. Are you Gram-positive or Gram-negative? (ou can make a good guess about this ever if you didn' see your cells)

Explanation / Answer

1 a) i. Crystal violet dissociates in aqueous solutions into CV+ and Cl-. The CV+ ions interacts with negatively charged components(peptidoglycan) of bacterial cells and stains the cells.

ii. Iodide interacts with CV+ and forms large complexes of crystal violet and iodine within the layers of the cell.

iii. Without Iodine the crystal violet stain may decolourized on ethanol wash.

b) Failure to add the iodine all cells will appear purple since the crystal violet stains both Gram-positive and Gram-negative cells, and the safranin is not likely to be seen over the darker crystal violet.

c) Gram-positives will be purple and gram-negatives will be colourless since they have been decolourized but not counterstained.

d) On reversal of crystal violet and safranin stains: Safranin will be washed out during decolourization, so bothe will be colourized by 'counterstain' crystal violet. If the safranin is not completely washed away, the darker crystal violet will cover the lighter safranin.

2. As mentioned above that the ability of Gram-positive cells to retain the crystal violet when an alcohol decolourization. This ability is enhanced with the use of the mordant iodine.

3. Epithelial cells do not have peptidoglycan. So they are gram-negative. As they do not retain the crystal violet so they do not stain.

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