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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. |