EXERCISE 3

Question 1

Why should you air dry smear before fixation?

Answer 1

Water has tendency to destroy specimen through boiling

Question 2

if smear is from agar culture, what to do?

if from liquid medium?

Answer 2

mix small amount with water then air dry

spread out thinly

Question 3

Heat fixation DONT SCORCH

how many times to pass through the flame?

Answer 3

2-3 times (film-side up)

Question 4

CHEMICAL fixation

how much alcohol to spread and how long?

Answer 4

95% for 1 minute

Question 5

Common substance that requires chemical fixation and why

Answer 5

Milk

bc it CURDLES during heat -> poor smear

Question 6

Purpose of fixation (3)

Answer 6

1. kill microogranisms (denatures proteins)
2. appear thicker and bigger for visibility
3. stickier to not be washed away and adhere to

Question 7

Kind vs Type of stain

Answer 7

Kind : Basic & Acidic
Type : Simple, Differential, & Structural

Question 8

1st kind of stain

Answer 8

Basic dye (cationic)

Question 9

Examples of basic dye (5)

Answer 9

1. methylene blue
2. basic fuschin
3. crystal violet
4. safranin
5. malachite green

Question 10

Opposite of basic dye and function

Answer 10

Acidic dye (anionic) to stain bg

Question 11

Examples of acidic dye

Answer 11

Nigrosin
Indian Ink

Question 12

SIMPLE STAINING

• how many stains used
• 2 kinds of stain used

Answer 12

2
basic/acidic

Question 13

DIFFERENTIAL STAINING

• how many stains used
• most common example

Answer 13

• more than 2 to differentiate between 2 specimen
• gram staining

Question 14

Gram stain
- based on what part of the cell
- procedure steps and time length

Answer 14

-cell wall composition

Crystal Violet (1 min)
Iodine (1 min)
Alcohol (20 secs)
Safranin (1-2 min)

Question 15

what are endospores (green)

endosporulation

Answer 15

resistant to heat, harsh chemicals, radiation, and stain

Question 16

What can stain endospores?

Answer 16

Malachite Green (endospores) then Safranin (vegetative cells)

Question 17

Why are dyes used on flagella?

Answer 17

to increase diameter

Question 18

example of stain used on flagella

and what color does it show

Answer 18

heavy metals

red to pink

Question 19

What do you call bacteria with 1 flagellum?

—- at the end of one side?
—- at both sides?

Answer 19

monotrichous

polar

amphitrichous

Question 20

what di you call bacteria with multiple flagella at…

—- one end
—- all around

Answer 20

lophotrichous

peritrichous

Question 21

capsule
-definition
-location
-composition

Answer 21

• gelatinous outer layer
  that is secreted by the cell
• surrounds and adheres to cell wall
• complex polysaccharides

Question 22

most difficult staining and why

Answer 22

capsule staining - capsules are water soluble and should not be heated

Question 23

stains used in capsule staining

Answer 23

crystal violet ( 1% aqueous)
copper sulfate (20% for decolorizing)

FAINT BLUE COLOUR

Question 24

other examples of MORDANT and DECOLORIZER

Answer 24

heat and water

Question 25

how can water be used as a decolorizer during endospore staining

Answer 25

malachite green is water-soluble

Question 26

common shape of endospores

Answer 26

spherical or elliptical

Question 27

What kind of staining does capsule staining use?

Answer 27

Both acidic and basic dye to emphasize color of capsule

Question 28

Does capsule staining use mordant?

Answer 28

No