Structural Staining (P)

Question 1

What happens in Schaeffer-Fulton’s method (of endospore staining)?

Answer 1

A primary stain (malachite green) is forced into the spore by steaming the bacterial emulsion

Question 2

What are the characteristics of malachite green?

Answer 2

1) It is water soluble

2) It has low affinity for cellular material

Question 3

What is the result from the characteristics of malachite green?

Answer 3

Vegetative cells may be decolorized with water

Question 4

What is the purpose of safranin?

Answer 4

It is applied to counterstain any cells which have been decolorized

Question 5

What are the results at the end of staining?

Answer 5

1) Vegetative cells will be pink

2) Endospores will be dark green

Question 6

What are the reagents used in endospore staining (Shaeffer-Fulton’s method)?

Answer 6

1) Malachite green
2) Water
3) Safranin

Question 7

What is the primary stain used in endospore staining (Schaeffer-Fulton’s method)?

Answer 7

Malachite green

Question 8

What is the decolorizing agent used in endospore staining (Schaeffer-Fulton’s method)?

Answer 8

Water

Question 9

What is the counterstain used in endospore staining (Schaeffer-Fulton’s method)?

Answer 9

Safranin

Question 10

What is the procedure of endospore staining (Schaeffer-Fulton’s method)?

Answer 10

1) Application of malachite green (primary stain)]
2) Application of heat (mordant)
3) Application of water (decolorizer)
4) Application of safranin (counterstain)

Question 11

What is the mordant used in endospore staining (Schaeffer-Fulton’s method)?

Answer 11

Heat

Question 12

What are the characteristics that should be considered in the differentiation of genera and species identification of bacteria?

Answer 12

1) Flagella (which is possessed by most motile bacteria)
2) Shape
3) Number
4) Position

Question 13

Can bacterial flagella be stained?

Answer 13

Yes

Question 14

How does flagella staining differ from staining of other bacterial structures?

Answer 14

It differs from staining other bacterial structures because flagella staining usually requires extraordinary care for the slides, stain, and cells

Question 15

What is the purpose of painstakingly preparing flagellar stains?

Answer 15

These are prepared to coat the surface of the flagella with dye or a metal such as silver

Question 16

What is the procedure of preparation of smear for flagellar staining (modified Ryu’s method)?

Answer 16

1) Take a slant of flagellated cell structure
2) Add 2 drops of sterile distilled H20
3) Incubate (20 mins)
4) Take a drop of suspension and add on slide kept in slanting position
5) Air dry the smear and no heat fixation treatment is given to the smear
6) Flood the air-dried smears with the staining solution for 1 - 5 mins
7) Wash the staining solution off in tap H20. After the smears have dried, examine them under the OIO of microscope. Cell bodies and flagella stain violet

Question 17

What is the resulting color of staining flagella?

Answer 17

Violet (cell bodies are also violet)

Question 18

What is the characteristic of capsules to reagents in simple staining?

Answer 18

Capsules stain very poorly with reagents used in simple staining

Question 19

Capsule stain can become what (depending on the method)?

Answer 19

A misnomer

Question 20

Why do capsule stain become a misnomer?

Answer 20

Because the capsule may or may not be stained

Question 21

What are (-) staining methods?

Answer 21

These contrasts a translucent, darker colored background

Question 22

What are the results of (-) staining methods?

Answer 22

1) Cells are stained

2) Capsule is unstained

Question 23

What are used to form the background in (-) staining methods?

Answer 23

1) India ink
2) Nigrosin
3) Congo red

Question 24

Compare india ink and nigrosin

Answer 24

India ink: difficult to obtain nowadays

Nigrosin: easily acquired

Question 25

What is required for a (+) capsule stain?

Answer 25

A mordant

Question 26

What is the purpose of mordant in a (+) capsule stain?

Answer 26

It precipitates the capsule

Question 27

What are the counterstains that can be used in capsular staining?

Answer 27

1) CV

2) Methylene blue

Question 28

What happens in counterstaining (in capsular staining)?

Answer 28

The bacterial cell wall takes up the dye

Question 29

Result for capsules against a dark bg?

Answer 29

Capsules appear colorless

Stained cells are present

Question 30

What is the procedure of congo red capsule staining?

Answer 30

1) Place a loop-full of Congo red on a slide
2) Mix a small amt of your organism into the drop of Congo red
- > spread the organism or dye suspension well on the slide
3) Let the slide thoroughly air dry
- > do not methanol fix!
4) Fix the dried slide with acid alcohol (for 15 secs)
5) Rinse with distilled H20 and cover the slide with acid fuchsin for 1 - 5 mins
6) Rinse with H20 and allow to air dry
7) Examine the slide under oil immersion
- > cells stain red / pink, and the capsules appear as colorless halos against a dark blue bg