Flashcards in LAB: Differential stains Deck (15):
1
Gram Stain Procedure
- Prepare a smear of the bacteria
- Apply Crystal violet. Sit for 1 minute and rinse with distilled water
- Apply Gram's Iodine. Sit for 1 minute and rinse with distilled water
- Rinse gently with ethyl alcohol. Do not over rinse - rinse for few seconds. Rinse with distilled water for 10-15 seconds
- Apply Safranin. Sit for 1 minute and rinse with distilled water
- Blot dry with bibulous paper
- Observe on high power
- Sketch and label Gram + control and Gram - control
2
Gram Stain Procedure cont...
-
Purple indicates Gram-positive (Staphylococcus; cocci in clusters)
-
Pink indicates Gram-negative (E. coli; individual rods)
3
Gram-Positive Composition of cell wall
- thick peptidoglycan
- no outer membrane
- stained purple
- cells are stained
- uncolored around the cell is the capsule
- cells are stained
- uncolored around the cell is the capsule
4
Gram-Negative Composition of Cell Wall
- thin peptidoglycan
- outer membrane (highly negative)
- stained pink
5
Capsule Stain
- background purple stained
- capsule is halo-like
6
Importance of capsule
- Hides bacteria from immune system; gives some bacteria its virulence
- adhesion
- made of polysaccharides
- thick rigid layer
- thick rigid layer
7
Endospore Stain
- differentiate between spores and vegetative cells
- pink stained cells = vegetative cells
- light greened stained cells = endospores
-
Schaeffer-Fulton endospore slide
8
Central location
- Central endospores are located in the middle of the vegetative cell
9
Terminal location
- Terminal endospores are located at the end of the vegetative cell
10
Examples of bacteria that forms endospores
- Bacillus
- Clostridium
11
Differential Stains
- use 2 different dyes
- differentiate between cell types or cell structures
12
Primary Stain
- Crystal Violet - stains all cells purple
13
Mordant
-
Gram's Iodine - fixes crystal violet into cell wall of G+ cells
14
Decolorizer
-
Ethyl Alcohol - removes crystal violet from G- cells
15
Counterstain