Chocolate agar
Tryptic Soy Agar supplemented with 5% sheep blood
DESCRIPTION
Highly nutritive medium used for the isolation
and cultivation of various fastidious
microorganisms.
The medium is also used to determine the hemolytic capacity of
bacteria.



COMPOSITION
Nutrient agar composition:
Beef extract 10g, Peptone 10g, NaCl 5g, Agar 20g.
Add distilled water ad 1000ml; adjust pH to 7.2 at 25 ºC. Sterilize by autoclaving at 121
ºC for 15 minutes.


PROCEDURE

Melt nutrient agar base, cool to 45-50 ºC then add 5% sterile, defibrinated blood
(sheep, bovine or horse origin). For
Haemophilus spp. and Neisseria spp.  prepare
chocolate agar by adding blood at higher temperature (70-80 ºC).


NOTE
Some bacteria ha
ve the ability to produce toxins that lyse erythrocytes from the culture
substrate (blood agar), resulting a greenish or clear halo surrounding the colonies
(partial or complete hemolysis). The red cells are easily destroyed by other factors
than bacterial activity, like pH value
, salinity of the inoculum, improper storage
conditions of the culture medium like freezing, heating above 50 ºC, or adding acidic
or basic supplements.


QUALITY CONTROL
Following microorganisms should have a good growth at 35 ºC in 24 hours:
Neisseria meningitidis ATCC 13090,
Streptococcus pneumoniae ATCC 6305,
Streptococcus pyogenes ATCC 19615,
Bacillus cereus ATCC 11778.

Following microorganisms should have a good growth at 28 ºC in 3 days:
Candida albicans ATCC 10231,
Aspergillus niger ATCC 16404.
Blood agar
Sheep blood agar
Chocolate agar
(c) Costin Stoica
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Sheep blood agar medium destroyed by freezing
A - alpha-hemolysis, B - beta-hemolysis, C - false
hemolysis produced by acidic inoculum