1. The Gram stain Thin smear/heat fix Gram stain:
a. Flood slide with crystal violet and let stain for 1 minute.
b. Drain off crystal violet and rinse off with distilled water; flood slide with Gram's iodine for 1 minute.
c. Rinse off Gram's iodine with distilled water.
d. Hold the slide on an angle (preferably with a clothes pin) and drop 95% ethyl alcohol onto it until the alcohol leaving the slide no longer has a purple tint; be sure to drop the alcohol onto the upper portion of the slide so that the smears are subjected to uniform decolorization. Be careful not to
"decolorize" dye from the clothes pin!!
e. Rinse with distilled water and flood the slide with safranin and let stain for minutes.
f. Rinse with distilled water and blot dry with bibulous paper.
Gram
positive
Gram
negative

2. Gram-positive cell walls Gram-negative cell walls
The characteristic compound found in all true bacterial cell walls
is peptidoglycan. The amount of PPG is among one of the
differences between the GP and GN cell walls.
Gram-positive cell walls Gram-negative cell walls
Thick peptidoglycan
90% peptidoglycan
Teichoic acids
1 layer
Not many polysaccharides
In acid-fast cells, contains mycolic acid
Thin peptidoglycan
5-10% peptidoglycan
No teichoic acids
3 layers
Outer membrane has lipids, polysaccharides
No acid- fast cells (mycolic acid)

5. Examples:
Examples of gram-negative bacteria: Spirochetes (spiral-shaped) - causes syphilis, lyme disease Neisseria (cocci) - causes meningococcus, gonorrhea
Our Favorite: E. coli!!

6. Six common gram-positive bacteria that infect humans and their shapes follow: Streptococcus (cocci) Staphylococcus (cocci) Bacillus (bacilli, protective spore) - causes anthrax and gastroenteritis Clostridium (bacilli, protective spore) - causes botulism, tetanus, gas gangrene, and pseudomembranous colitis Corynebacterium (bacilli, no protective spore) - causes diphtheria Listeria (bacilli, no protective spore) - causes meningitis