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Blood Agar is a general-purpose enriched medium used to grow fastidious organisms and to differentiate bacteria based on their hemolytic properties. It is a highly nutritious medium generally used as a basal medium for preparing blood agar by supplementation with blood. It is typically Tryptic Soy Agar or Columbia Agar base with 5% Sheep blood.
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Composition of Blood Agar
Composition per liter:
Agar = 15.0 gm
Beef extract = 10.0 gm
Peptone = 10.0 gm
Sodium Chloride (NaCl) = 5.0 gm
Sheep blood, defibrinated = 50.0 mL
pH 7.3 ± 0.2 at 25°C
Principle of Blood Agar
Beef extract and Peptone provide carbon, nitrogen, amino acids and vitamins. Sodium chloride helps in maintaining the osmotic equilibrium of the medium. Agar is a solidifying agent. The addition of blood makes the medium more nutritious by providing additional growth factors required by fastidious organisms. It also helps in visualizing the hemolytic reactions. However, hemolytic reactions depend on the animal blood used.
Sheep blood gives the best results for Group A Streptococci. But sheep blood fails to support the growth of Haemophilus haemolyticus since sheep blood is deficient in pyridine nucleotides. However when horse blood is used H. haemolyticus colonies produce hemolysis and mimic Streptococcus pyogenes.
Preparation of Blood Agar
- Add the above components (40 gm), except sheep blood, to distilled/deionized water and bring volume to 950.0 mL.
- Mix thoroughly.
- Heat with frequent agitation and boil for 1 min to completely dissolve.
- Autoclave for 15 min at 15 psi pressure at 121°C.
- Cool to 45°- 50°C.
- Aseptically add 50.0 mL of sterile, defibrinated sheep blood.
- Mix thoroughly and pour into sterile Petri dishes.
- It is used for the isolation and cultivation of Streptococci, Neisseria and other fastidious microorganisms.
- It is used to differentiate bacteria on the basis of their hemolytic properties (β-hemolysis, α-hemolysis, and γ-hemolysis (or non-hemolytic)).
- To determine the salinity range of marine Flavobacteria.
- It can also be used for the preparation of Salmonella Typhi antigens.
- It is recommended by APHA and Standard Methods for the testing of food samples.