ATCC medium: 2192 CEM medium
Eugon Agar (BD 258910)..................45.0 g
IsoVitalex Enrichment (BD 211876).......10.0 ml
Horse blood, defibrinated...............50.0 ml
Horse blood, laked......................50.0 ml
Distilled water........................900.0 ml
Suspend Eugon Agar in 900.0 ml of distilled water. Boil to dissolve the agar. Autoclave at 121C for 15 minutes.
Adjust to pH 7.0 +/- 0.2. Cool to 75-80C in water bath. Add defibrinated horse blood to this hot agar solution.
Adding blood to hot(75-80C) medium causes the blood to "chocolatise" (turn dark brown).
Cool to 50-55C in water bath. Add laked horse blood to cooled medium.
To "lake" horse blood, aseptically measure blood and pour into a sterile glass bottle. Place the bottle in a freezer until frozen.
Thaw the blood at room temperature. Repeat the freezing/thawing process 3 more times to disrupt and lyse the blood cells.