Download Slide 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 2-3
minutes.
f. Rinse with distilled water and blot dry with
bibulous paper.
Gram
positive
Gram
negative
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
walls
• Thick
peptidoglycan
• 90% peptidoglycan
• Teichoic acids
• 1 layer
• Not many
polysaccharides
• In acid-fast cells,
contains mycolic
Gram-negative cell
•
•
•
•
•
Thin peptidoglycan
5-10% peptidoglycan
No teichoic acids
3 layers
Outer membrane has
lipids, polysaccharides
• No acid- fast cells
(mycolic acid)
Examples:
• Examples of gram-negative bacteria:
Spirochetes (spiral-shaped) - causes
syphilis, lyme disease
Neisseria (cocci) - causes
meningococcus, gonorrhea
– Our Favorite: E. coli!!
• 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