bacterial smear preparation and simple stain Flashcards
This is done when basic stain is applied
Simple staining
What type of stain do we use in simple staining?
Basic stain
These helps us identify the bacteria under the microscope after applying staining reagent to the prepared smear
- Size
- Shape
- Arrangement
It is done after air drying the bacteria in the glass slide in smear preparation
Heat Fix
What electric charge do stain carries which the surface and cytoplasm of bacteria attracts?
Positive electric charge
What electric charge do the surface or cytoplasm of bacteria carries which stain attracts?
Negative electric charge
Two types of sample in the
DIRECT SMEAR
- from SWABS
- Concentrated centrifuged specimens
What do we do to avoid destruction of cellular entities in direct smear preparations when using swab?
Rolling the swab across the slide
In preparing concentrated centrifuged specimens in a glass slide, what do we use to transfer the sample?
Plastic transfer pipette
Needs to be sterile
Three types of samples for SMEARS
- Culture smears (subculture smears)
- Blood culture bottles
- Solid media
How many slide do we use in preparing smears from blood culture bottles?
One slide only
It is used to puncture the top of the blood culture bottle
Safety venting subculture unit
It is used for placing a small drop of water in the squares of glass slides
(not sure with this question)
I-mL tuberculin syringe
with the needle removed
These are added first before smearing bacteria to the glass slide
- Sterile NSS
- Distilled water (H20)
Why do we rub the colony into the glass slide with a drop of H20 or distilled water in it?
for emulsification
or to emulsify
What should be the appearance of the glass slide after emulsifying it?
Turbid
Two types of culture
- Pure culture
- Mixed culture
Three types of tool used for implanting the bacteria into the glass slide
- Inoculating loop
- Inoculating needle
- Disposable Inoculating loop and needle
Two sizes for inoculating loop
- 10uL
- 1uL
It is commonly reffered to the use of a single stain to color a bacterial organism
Simple staining
How many stain do we use in simple staining?
1 stain only
Three common types of stains in simple staining
- Methylene blue
- Basic fuchsin
- Crystal violet.
Why do we use stain to color bacterial organism?
It is useful in determining basic morphology and the presence or absence of certain kinds of granules.
How many seconds/minutes is done in simple staining?
30 seconds to 2 minutes
depending on the affinity of the dye
It is where we place the glass slides for gram staining or acid-fast staining
Staining rack
What are the steps in gram staining?
- Primary stain
- Mordant
- Decolorizer
- Secondary stain/counterstain
Primary stain used in gram staining
crystal violet
Mordant used in gram staining
Gram’s Iodine
Decolorizer used in gram staining
Acetone alcohol
Secondary stain/counterstain used in gram staining
Safranin
Typical time for flooding a glass slide with primary stain in gram staining
1 minute
Typical time for flooding a glass slide with mordant in gram staining?
1 minute
Typical time for flooding a glass slide with decolorizer in gram staining
3 seconds or until the stain comes off
Typical time for flooding a glass slide with secondary stain in gram staining
30 seconds
What do we do between the steps performed in gram staining?
Wash off the slides with distilled water
What are the steps in acid-fast staining?
- Primary stain
- Mordant
- Decolorizer
- Secondary stain/counterstain
Two methods in acid-fast staining?
Hot and Cold method
What does the HOT method called in acid-fast staining?
Ziehl-Neelsen Method
What does the COLD method called in acid-fast staining?
Kinyoun Method
Primary stain used in acid-fast staining
Carbol Fuchsin
Mordant used in HOT method for acid-fast staining
Heat
Decolorizer used for HOT method in acid-fast staining
Acid alcohol
Counterstain used for HOT method in acid-fast staining
Methylene blue/Malachite green
Typical time for flooding a glass slide with primary stain in acid-fast staining?
2 minutes
Typical time for applying heat to the glass slide using alcohol lamp in acid-fast staining?
5 minutes
Typical time for flooding a glass slide with decolorizer in acid-fast staining?
3 seconds or until the stain comes off
Typical time for flooding a glass slide with secondary stain in acid-fast staining?
1 minute
What is the color of a gram POSITIVE bacteria after applying counterstain in gram staining?
Purple
What is the color of a gram POSITIVE bacteria after applying decolorizer in gram staining?
Purple
What is the color of a gram NEGATIVE bacteria after applying counterstain in gram staining?
Pink/Red
What is the color of a gram NEGATIVE bacteria after applying decolorizer in gram staining?
Colorless
What is the color of a gram POSITIVE bacteria after applying counterstain in acid-fast staining?
Red
What is the color of a gram NEGATIVE bacteria after applying counterstain in acid-fast staining?
Blue/Green
What is the color of a gram POSITIVE bacteria after applying decolorizer in acid-fast staining?
Red
What is the color of a gram NEGATIVE bacteria after applying decolorizer in acid-fast staining?
Colorless
What is the composition of the acid alcohol used in HOT method?
3% hydrochloric acid (HCL) + 95% ethanol
What is the composition of the acid alcohol used in COLD method?
3% sulfuric acid (H2SO4) or 0.5% hydrochloric acid (HCL) + 95% ethanol
The position of the inoculating straight wire in the alcohol lamp
Diagonal downward
Length of the inoculating straight wire
not more than 5cm
Composition of the inoculating straight wire
Made up of: Nichrome wire (or platinum)
We flame the inoculating straight wire until _____
Red hot
