Mesoplama florum:Culture media

ATCC 1161 Medium
Mix a base broth in a 1 liter 45mm cap bottle the following chemicals:

Autoclave at 121C for 20 minutes and cool to room temperture (broth) or 55C (Agar) on a 55C water bath.

Centrifuge 2 x 45 ml of Gibco 18180-059 liquid yeast extract at high speed to remove sediment. Pour only 45 ml into centrifuge bottles, eliminating as much as possible of the sediment in the bottom of the Gibco stock.

Thaw Sigma H-1138 heat inactivated horse serum. For agar, bring both horse serum and centrifuged yeast extract to 37C. Horse serum at 55C will sediment protein.

Assemble 500 ml bottle top 0.22&mu; filter on the 1 liter bottle. For agar, it is helpful to put the bottle on a foam insulating pad to delay cooling. Add by vacuum filtration:

Adjust pH to 7.4 with 1 M HCl or 1 M NaOH. Measured pH of as-made broth is 7.46, probably close enough. For cresol red containing solution, adjust pH to 7.8

Phenol red 2% solution must be neutralized with sufficient NaOH to convert it to the sodium salt. The phenol or cresol red amount can be adjusted to control the depth of color to taste.

SP-4 (ATCC 988) Medium
Mix a base broth containing:

Adjust pH to 7.5 and autoclave at 121C for 20 minutes.

Centrifuge 2 x 17.5 ml of liquid yeast extract (Gibco 18180-059) at high speed to remove sediment.

Mix 2 g of Yeastolate (Difco 5577) in 100 ml DI water.

Sterile filter into the cooled medium: