predict the effect of the following mistakes made when performing a gram stain


What is a common mistake which occurs when performing a Gram stain?

In this study, we present a review of over 6,000 Gram-stains and establish an error rate of around 3%, with the most common reason for error being an over-decolourisation step resulting in organisms that should be Gram-positive appearing as Gram-negative.

What would happen if you made an error and reversed the crystal violet and safranin stain?

If there is a reversal of crystal violet and safranin stains, then the cross-link between iodine and crystal violet won’t happen, and during decolorization, the safranin will fade away. Ans also, as crystal violet stain is used in the last, it will make all cells purple.

What is a potential problem with the Gram stain technique?

The Potential problems or errors in Gram staining techniques. • Decolorizing too much or little. • Washing too much of the crystal violet off with water.

What does Gram staining effect?

A Gram stain is a test that checks for bacteria at the site of a suspected infection or in certain body fluids, such as blood or urine. These sites include the throat, lungs, and genitals, and in skin wounds. There are two main categories of bacterial infections: Gram-positive and Gram-negative.

Why would a gram stain be wrong?

Positive Gram stains with negative culture results could be due to fastidious or nonviable organisms and failure to order appropriate testing, such as anaerobe cultures. False-negative Gram stains could occur due to inadequate specimen or smear preparation or failure to examine an adequate number of fields.

Which step in the gram stain procedure is most prone to error if done incorrectly How might that step affect the end result?

Which step in the gram stain procedure is most prone to error? If done incorrectly, how might that step affect the end result? Too much decolorizer = g+ lose initial stain.

What would happen if you use safranin first and crystal violet second?

If you reverse the staining procedure that is using safranin first, this will cause all the bacteria to remain red and crystal violet applied later on may cause the gram-negative bacteria to become violet in color and the gram -positive bacteria will remain red.

What would be the failure to apply the safranin stain?

3) Failure to apply the safranin. You would not be able to see the Gram-negative cells. The Gram-positive would be the only cells that are colored and therefore visible. … Both Crystal Violet and Safranin are basic stains and may be used to do simple stains on gram positive and gram-negative cells.

What would happen if you left out the application of safranin?

What would happen if you left out the safranin when performing the Gram stain? … You must heat fix your smear before you perform the acid fast staining technique.

Why might gram-negative incorrectly stain purple?

When the counter stain of safranin is added the decolorized gram-negative bacteria stain pink. In contrast, the solvent dehydrates the thicker gram-positive cell walls, closing the pores as the cell wall shrinks. As a result, the violet-iodine complex is retained and the cells remain purple.

Which step in the Gram stain is most likely to cause poor results if done incorrectly and why?

The decolorization of the cells is the most “operator-dependent” step of the process and the one that is most likely to be performed incorrectly. Rinse with water to stop decolorization. Rinse the slide with a counterstain (safranin or carbol fuchsin) which stains all cells red.

How does culture age affect the results of a Gram stain?

how does culture age affect the results of a gram stain? if the cultures are more than 16-18 hours old, then Gram-positive cultures can stain gram-variable or gram-negative and give false results. … Thick smears can trap stain and falsely make cells appear gram positive.

What are the disadvantages of Gram staining?

Disadvantages: Some bacteria are Gram stain variable (positive or negative results) Some bacteria are resistant to Gram stain (i.e. acid-fast bacteria) False results may occur if over-decolorized.

What is the theory about the mechanism of the Gram stain reaction?

What is the current theory about the mechanism of the gram stain reaction? The gram stain is a basic differential stain used to determine if a bacterial cell is gram positive or negative. Gram positive cells have a thick peptidoglycan layer that will trap the crystal violet iodine crystals and appear purple.

What might cause a gram-negative to be seen as a gram positive?

If the decolorizing agent is applied on the cell for too long time, the Gram-positive organisms to appear Gram-negative. Under-decolorization occurs when the alcohol is not left on long enough to wash out the CV-I complex from the Gram-negative cells, resulting in Gram-negative bacteria to appear Gram-positive.

Why is it important to do a Gram stain correctly?

The main benefit of a gram stain is that it helps your doctor learn if you have a bacterial infection, and it determines what type of bacteria are causing it. This can help your doctor determine an effective treatment plan.

When performing any staining technique What problems can occur if the stain is left on the bacterial smear too long?

1. When performing any staining technique, what problems can occur if the stain is left on the bacterial smear too long? In a simple stain, probably the worst consequence is that the cell may appear larger than it really is.

What is negative staining in microbiology?

Negative staining employs the use of an acidic stain and, due to repulsion between the negative charges of the stain and the bacterial surface, the dye will not penetrate the cell. In negative staining, the results yield a clear cell with a dark background.

What is the correct order of staining reagents in Gram staining?

Explanation: Gram staining is a type of differential staining. In this process the fixed bacterial smear is subjected to the following staining reagents in the order listed: crystal violet, iodine solution, alcohol (decolorizing agent), and safranin. 3.

Why do we use positive and negative controls in the Gram stain quizlet Labster?

The positive and negative controls are bacterial smears we use to test if the Gram stain was performed properly. If the positive and negative controls are not as expected, you can not trust that the stain was performed properly.

What is the most critical step in the Gram staining procedure?

The critical step of the Gram staining procedure is the decolorization step. Hold the slide in a tilted downward position and allow the decolorizer to flow over the smear.

What happens if you stain with safranin before crystal violet?

Since the safranin is lighter than crystal violet, it does not disrupt the purple coloration in Gram positive cells. However, the decolorized Gram negative cells are stained red.

What would happen if you switched the order of the crystal violet and the safranin?

What would you anticipate seeing if you accidentally switched crystal violet and safranin while performing a Gram stain? All bacteria would appear purple.

How might the results of the gram stain be impacted if the primary and secondary stain were switched?

If the dyes are switched in a gram stain the a gram positive cell will appear pink and a gram negative will appear purple. … Gram negative stains will not be decolorized and will retain the purple color from crystal violet.

What happens if you do Gram stain backwards?

Do NOT decolorize for a full minute!

If the decolorizer is left on too long, even gram positive cells will lose the crystal violet and will stain red.

What happens if you over Decolorize in the Gram stain procedure?

Over-decolorizing will lead to an erroneous result where gram-positive cells may stain pink to red indicating a gram-negative result, and under-decolorizing will lead to an erroneous result where gram-negative cells may appear blue to purple indicating a gram-positive result.

What is safranin in Gram staining?

The safranin is also used as a counter-stain in Gram’s staining. In Gram’s staining, the safranin directly stains the bacteria that has been decolorized. With safranin staining, the gram-negative bacteria can be easily distinguished from gram-positive bacteria.

What would be the consequence if you forgot to apply safranin in Gram staining technique?

Safranin is used to color the Gram-negative bacteria that have been decolorized in the previous step. At the beginning of this video, you will note that the Gram-negative cells have no color. If the safranin step is omitted, then the Gram-negative cells will be colorless and difficult to see.

What outcome would be expected if you forgot the crystal violet step while performing a Gram stain?

What outcome could be expected if you forgot the crystal violet step while performing a Gram stain? Gram-positive and gram- negative organisms would stain pink. If you forget to use the decolorizer, which cell type(s) will appear purple?

What are the common errors in staining methods?

The rank of errors is:
  • Over decolourisation.
  • Mixed cultures.
  • Misread stains.
  • Aged subcultures.
  • Disorganisation.
  • Inadequate fixation and Insufficient culture.

Why do Gram negatives stain pink?

Gram-negative cells have a thinner peptidoglycan layer that allows the crystal violet to wash out on addition of ethanol. They are stained pink or red by the counterstain, commonly safranin or fuchsine. … This gives rise to gram-variable and gram-indeterminate groups.

Which of these factors will affect your gram stain results?

The many variables that can affect this stain are age of the culture, amount of decolorizer used, the time of decolorization, the type of organism (acid-fast bacteria and spores do not stain well), thickness of the smear, and the general care of the stainer.

Gram Staining Procedure Animation Microbiology – Principle, Procedure, Interpretation

Gram Staining

How to prepare the perfect Gram stain

Performing the Gram Stain

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