EXERCISE 3 Flashcards
Why should you air dry smear before fixation?
Water has tendency to destroy specimen through boiling
if smear is from agar culture, what to do?
if from liquid medium?
mix small amount with water then air dry
spread out thinly
Heat fixation DONT SCORCH
how many times to pass through the flame?
2-3 times (film-side up)
CHEMICAL fixation
how much alcohol to spread and how long?
95% for 1 minute
Common substance that requires chemical fixation and why
Milk
bc it CURDLES during heat -> poor smear
Purpose of fixation (3)
- kill microogranisms (denatures proteins)
- appear thicker and bigger for visibility
- stickier to not be washed away and adhere to
Kind vs Type of stain
Kind : Basic & Acidic
Type : Simple, Differential, & Structural
1st kind of stain
Basic dye (cationic)
Examples of basic dye (5)
- methylene blue
- basic fuschin
- crystal violet
- safranin
- malachite green
Opposite of basic dye and function
Acidic dye (anionic) to stain bg
Examples of acidic dye
Nigrosin
Indian Ink
SIMPLE STAINING
- how many stains used
- 2 kinds of stain used
2
basic/acidic
DIFFERENTIAL STAINING
- how many stains used
- most common example
- more than 2 to differentiate between 2 specimen
- gram staining
Gram stain
- based on what part of the cell
- procedure steps and time length
-cell wall composition
Crystal Violet (1 min)
Iodine (1 min)
Alcohol (20 secs)
Safranin (1-2 min)
what are endospores (green)
endosporulation
resistant to heat, harsh chemicals, radiation, and stain
What can stain endospores?
Malachite Green (endospores) then Safranin (vegetative cells)
Why are dyes used on flagella?
to increase diameter
example of stain used on flagella
and what color does it show
heavy metals
red to pink
What do you call bacteria with 1 flagellum?
—- at the end of one side?
—- at both sides?
monotrichous
polar
amphitrichous
what di you call bacteria with multiple flagella at…
—- one end
—- all around
lophotrichous
peritrichous
capsule
-definition
-location
-composition
- gelatinous outer layer
that is secreted by the cell - surrounds and adheres to cell wall
- complex polysaccharides
most difficult staining and why
capsule staining - capsules are water soluble and should not be heated
stains used in capsule staining
crystal violet ( 1% aqueous)
copper sulfate (20% for decolorizing)
FAINT BLUE COLOUR
other examples of MORDANT and DECOLORIZER
heat and water
how can water be used as a decolorizer during endospore staining
malachite green is water-soluble
common shape of endospores
spherical or elliptical
What kind of staining does capsule staining use?
Both acidic and basic dye to emphasize color of capsule
Does capsule staining use mordant?
No
