Apply mordant (Gram's iodine). The iodine combines with the crystal violet in the cell to form a crystal violet-iodine complex (CV-I).

Step 2 (two)in the staining technique.

Apply decolorizing agent (ethanol or ethanol-acetone). The primary stain is washed out of some bacteria (decolorized), whereas others are unaffected.

Step 3 (three) in the staining technique.

Apply counterstain (safranin). This basic dye stains the decolorized bacteria red.

Step 4 (four) in the staining technique.

Bacterial cell walls are complex lattice structures composed of layers of?

Peptidoglycans and other compounds.

What do you wash the crystal violet off with when Gram staining?

Distilled water from a wash bottle

What do you wash the Gram's iodine off with when gram staining?

Distilled water from a wash bottle

After decolorizing with 95% ethanol, what do you do next?

Immediately wash gently with distilled water.

What do you do after the safranin has been on for 30 seconds?

Wash the slide with distilled water, and blot it dry with a paper towel or absorbent paper.

What are some common sources of gram-staining errors?

The loop was too hot. Excessive heat was applied during the heat-fixing. The decolorizing agent (ethanol) was left on the smear too long. The culture was too old. The smear was too thick.

Why will very thick smears give inaccurate results?

Because they are hard to decolorize correctly.

Why do you need to immediately wash with distilled water after decolorizing?

Because it will continue to decolorize.

Why do gram-positive cells more than 24 hours old stain gram-negative?

Because the cell wall begins to deteriorate and change.

Can iodine be added before the primary stain in a Gram stain?

No, it means to complex with crystal violet.

Which step can you omit without affecting determination of the gram reaction?

Safranin; but you would only see gram-positive.

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A differential stain that enables you to classify bacteria as either gram-positive or gram-negative. It is used for identifying and classifying bacteria.

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Primary stain (Crystal violet). All bacteria are stained purple by this basic dye.

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Micro Lab Exercise 7

Question	Answer
A differential stain that enables you to classify bacteria as either gram-positive or gram-negative. It is used for identifying and classifying bacteria.	Gram stain
Primary stain (Crystal violet). All bacteria are stained purple by this basic dye.	Step 1 (one) in the staining technique.
Apply mordant (Gram's iodine). The iodine combines with the crystal violet in the cell to form a crystal violet-iodine complex (CV-I).	Step 2 (two)in the staining technique.
Apply decolorizing agent (ethanol or ethanol-acetone). The primary stain is washed out of some bacteria (decolorized), whereas others are unaffected.	Step 3 (three) in the staining technique.
Apply counterstain (safranin). This basic dye stains the decolorized bacteria red.	Step 4 (four) in the staining technique.
Bacteria that decolorize easily are referred to as?	Gram-negative
Bacteria that decolorize slowly and retain the primary stain are called?	Gram-positive
Bacterial cell walls are complex lattice structures composed of layers of?	Peptidoglycans and other compounds.
When performing a Gram stain, how long should the crystal violet stay on the slide?	30 seconds
When performing a Gram stain, how long should the Gram's iodine stay on the slide?	10 seconds
What do you wash the crystal violet off with when Gram staining?	Distilled water from a wash bottle
What do you wash the Gram's iodine off with when gram staining?	Distilled water from a wash bottle
After washing the grams iodine off, you decolorize it with 95% ethanol. Let the alcohol run through the smear for 10 to 20 seconds.	True
After decolorizing with 95% ethanol, what do you do next?	Immediately wash gently with distilled water.
What do you do after you gently wash with distilled water?	Add safranin for 30 seconds.
What do you do after the safranin has been on for 30 seconds?	Wash the slide with distilled water, and blot it dry with a paper towel or absorbent paper.
What are some common sources of gram-staining errors?	The loop was too hot. Excessive heat was applied during the heat-fixing. The decolorizing agent (ethanol) was left on the smear too long. The culture was too old. The smear was too thick.
Why will very thick smears give inaccurate results?	Because they are hard to decolorize correctly.
Why do you need to immediately wash with distilled water after decolorizing?	Because it will continue to decolorize.
What color will a gram-negative cell stain?	Pink
What color will a gram-positive cell stain?	Purple
Why do gram-positive cells more than 24 hours old stain gram-negative?	Because the cell wall begins to deteriorate and change.
Can iodine be added before the primary stain in a Gram stain?	No, it means to complex with crystal violet.
Which step can you omit without affecting determination of the gram reaction?	Safranin; but you would only see gram-positive.

Created by: KamiSeago

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