Worksheet: Staining techniques (4 pts) 1. Predict the effect of the following mi
ID: 142936 • Letter: W
Question
Worksheet: Staining techniques (4 pts) 1. Predict the effect of the following mistakes on the appearance of both Gram-positive and Gram-negative bacteria. In other words, when you look through the microscope at your bacteria, what would you see? Conside each mistake to have occurred independently of the others. Gram Positive A. Failure to add Gram's iodine: B. Skipping the decolorizer step C. Decolorizing for 1 minute D. Failure to add safranin E. Staining with safranin 1t and crystal violet 2 F. Failure to heat fix the slide G. Using acid-alcohol instead of acetone-alcohol during the decolorizing step. 2. What about the structure of a Gram-positive organism's cell wal alows it to stain purple and w Gram-negative organism's cell wall causes it to stain red?Explanation / Answer
A.
Gram +: CV-I complex won't form in this case , crystal violet stain won't be retained; it will rather appear pink because of counterstain
G negative :- no effect
Gram positive would look like Gram negative -; basic purpose of iodine is to increase affinity for peptidoglycan layer; if you don't' have iodine, it is likely that you will over decolorize and then cells will retain color of counterstain.
B.In this case both G+ and G- cells will look purple; Safranin won't be easy to see over the crystal violet in the Gram negative cells.
D.Gram positive +: no effect
Gram negative -: invisible gram negative cells ; will be colorless.
E.crystal violet stains both gram positive and gram negative cells. it's just that gram negative cells don't retain the crystal violet stain because alcohol strips the outer membrane of G- cells .
Safranin can then successfully be held by Gram negative cells.
F.Both the bacteria are likely to get washed away during the staining procedure.
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