1. Basal medium

   Special peptone	20 g
   Yeast extract	2 g
   Mannitol	20 g
   Pyruvic acid (Na salt)	2 g
   NaCl	1 g
   MgSO4.7H2O (10 mg/ml)	1 ml
   Agar	12 g
   Distilled water	756 ml

   
   
2. Irgasan (Ciba-Geigy) solution

   Irgasan 0.40% in 95% ethanol

   1 ml

   
   May be stored at -20°C up to 4 weeks.
   
   

    

3. Desoxycholate solution

   Sodium desoxycholate	0.5 g
   Distilled water	200 ml

   Bring to boil with stirring; cool to 50-55°C.

    

4. Sodium hydroxide, [5 N] 1 ml
5. Neutral red, [3 mg/ml] 10 ml
6. Crystal violet, [0.1 mg/ml] 10 ml
7. Cefsulodin (Abbott Labs), [1.5 mg/ml] 10 ml
8. Novobiocin, [0.25 mg/ml] 10 ml
9. Strontium chloride, [10%; filter-sterilized] 10 ml

May be stored at -70°C. Thaw to room temperature just before use.

Preparation: Add ingredients for solution A (basal medium) to water and bring to boil with stirring. Cool to about 80°C (10 min in 50°C water bath). Add solution B (Irgasan) and mix well. Cool to 50-55°C. Add solution C (desoxycholate); solution should remain clear. Add solutions D through H. Slowly add solution I with stirring. Adjust pH to 7.4 with 5 N NaOH. Dispense 15-20 ml into each petri dish. Commercially prepared dehydrated Yersinia selective agar (Difco) with supplements may be substituted. Follow manufacturer's instructions for preparation.

Hypertext updated by rim/cjm 2005-OCT-21