Quick Answer: What Can Go Wrong In Gram Staining?

Which step in the Gram stain is most prone to error?

This step washes away unbound crystal violet, leaving Gram-positive organisms stained purple with Gram-negative organisms colorless.

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..

What happens if you Decolorize a Gram stain too long?

Do NOT decolorize for a full minute! The decolorizer should stay on the slide for no more than 15 seconds! If the decolorizer is left on too long, even gram positive cells will lose the crystal violet and will stain red.

What is the most important step in Gram staining?

The thickness of the smear used in the Gram stain will affect the result of the stain. The step that is most crucial in effecting the outcome of the stain is the decolorizing step.

Is a positive control properly stained?

Positive control: Staphylococcus aureus (should stain blue to dark purple/gram-positive cocci) Negative control: Escherichia coli (should stain pink to red/gram-negative rods)

What is the difference between a gram positive and gram negative bacteria?

Gram positive bacteria have a thick peptidoglycan layer and no outer lipid membrane whilst Gram negative bacteria have a thin peptidoglycan layer and have an outer lipid membrane.

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.

Are gram negative or gram positive bacteria harder to kill?

The major difference is the outer lipid membrane. It’s difficult to penetrate, which gives gram-negative bacteria extra protection. Gram-positive bacteria don’t have this feature. Because of this difference, gram-negative bacteria are harder to kill.

What color is gram negative bacteria?

RedGram negative organisms are Red. Hint; Keep your P’s together; Purple is Positive. Gram stains are never pink they are red or purple so you don’t destroy the rule; keep your P’s together. In microbiology bacteria have been grouped based on their shape and Gram stain reaction.

What happens if you don’t heat fix a Gram stain?

During the Gram stain you wash the bacteria several times with water and with alcohol and pat the slide dry at the end. Thus, if you do not heat fix, it is extremely likely that you will wash off your bacteria, leaving little to none to be seen under the microscope.

Why is iodine used in Gram staining?

When iodine is applied, decolorizing time for all cells is longer than without iodine. Thus, iodine penetrates the cell and serves to form a dye-iodine precipitate, and since all cells are less permeable for the dye-iodine toward decolorizing agents, slower removal of it results.

What is the importance of Gram staining?

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.

Why 95 Ethanol is used in Gram staining?

Remel Gram Decolorizer (95% Ethyl Alcohol) is a reagent recommended for use in qualitative procedures to differentiate gram-negative from gram- positive organisms. The primary stain, crystal violet, is a basic dye which rapidly permeates the cell wall of all bacteria, staining the protoplast purple.

What happens if you forget the Decolorizer in a Gram stain?

95% ethanol, because it removes the primary stain (crystal violet) from Gram-negative cells. If missed, then the bacteria would remain purple and give a false positive result.

Is E coli Gram positive or negative?

(E. coli) The gram-negative bacterium Escherichia coli is the most numerous aerobic commensal inhabitant of the large intestine. Certain strains cause diarrhea, and all can cause infection when they invade sterile sites (eg, the urinary tract).