Lecture 3B Microscopy and Staining Flashcards
Preparation of Specimens for Light Microscopy
- Smear
- Fixing
- Staining
Smear: Spread a __ of material containing microorganisms over the slide surface. Allow to __
- thin film
- air dry
Fixing: Process that __ microorganisms and __ them to a microscope slide. Fixing __ and __distortion of cells.
- kills
- attaches
- preserves
- minimizes
Staining: Coloring microorganisms with a dye that emphasizes certain structures. Before staining a sample, it must be __. Stains are __composed of __
- fixed
- salts
- 2 components: positive ion (cation) and negative ion (anion)
The colored ion in staining is called the __
chromophore
Two types of dyes
- basic dyes
- acidic dyes
A type of dye
- Chromophore is in __ ions.
Most commonly used dyes.
- Bacteria are slightly __ charged at pH 7, therefore they stain with basic dyes.
- Give examples
BASIC DYE
- positive
- negatively
- crystal violet, methylene blue, safranin (red/pink)
A type of dye
- Color is in __ ions.
- Stain the background: __
- Bacteria __ stain with acidic dyes.
- Used to observe __
- Minimal distortion because heat fixing is not necessary and dye is not taken up by cells.
- Give examples
ACIDIC DYE
- negative
- negative staining
- do not
- cell shape, size, and capsules
Examples:
- eosin (yellow)
- nigrosin (black/dark gray)
- india ink (black/dark gray)
cell wall __ the acidic dye
repels
Types of staining methods
- Simple Stains
- Differential Stains
a. Gram Stain
b. Acid-Fast Stain (Ziehl-Nielsen Stain) - Special Stains
a. Endospore Stain
b. Capsule Stain
c. Flagella Stain
A type of staining method
- Aqueous or alcohol solution of a single basic dye.
- Primary purpose is to stain the entire organism to view cell shape and basic structures.
SIMPLE STAINS
Procedure of simple staining:
- stain is applied for a certain time, then washed off
- slide is dyed and examined
A type of staining method
- React differently to different types of bacteria.
- Can be used to distinguish among different groups of bacteria.
Differential stains
two important differential stains used in microbiology
- gram stain
- acid fast stain (Ziehl-Nielsen Stain)
Who developed gram stain?
Hans Gram (1884), a Danish microbiologist