ATCC medium: 1161 Mycoplasma medium (ATCC 243) with sucrose
Heart Infusion Broth (BD 238400).. 17.5 g
Agar, Noble (BD 214230)........... 10.0 g
Horse serum*..................... 200.0 ml
Yeast extract solution (15%)..... 150.0 ml
Sucrose........................... 40.0 g
Distilled deionized water........ 650.0 ml
Combine heart infusion and 450 ml of water. Autoclave at 121C for 15 minutes. Add sucrose to 200 ml of water. Filter-sterilize. Aseptically add the horse serum, yeast extract solution, and sterile sucrose to the cooled heart infusion solution. Commercial yeast extract solution is sterile and need not be filtered. Final pH 7.4 +/- 0.2.
Dispense as desired. *Horse serum should be heat-inactivated at 56C for 30 minutes.
For solid medium, add 15 g of agar to the heart infusion solution, boil, and autoclave at 121C for 15 minutes. Cool this solution to 50-55C and warm the remaining solutions to 50-55C before combining.