Medium data

Search for medium no. = [144].

Unless otherwise stated, sterilize media by autoclaving at 121°C for 15 min.

144 MODIFIED NOS MEDIUM

Heart infusion broth (BD-Difco)	`1.25`	g
Trypticase peptone (BD-BBL)	`1.0`	g
Yeast extract (BD-BBL)	`0.25`	g
Sodium thioglycolate	`0.05`	g
L-Cysteine·HCl·H₂O	`0.1`	g
L-Asparagine	`0.25`	g
Glucose	`0.2`	g
0.1% Resazurin solution	`0.1`	ml
Distilled water	`94.0`	ml

Combine the above ingredients and bring to a boil in a round bottom flask while flushing with O₂-free N₂-H₂-CO₂ (85:10:5) gas mixture. Stopper anaerobically, wire or seal closed and autoclave at 121C for 20 min. After autoclaving, allow to cool, open without admitting oxygen and while flushing with gas, add the following ingredients which have been filter-sterilized ahead of time:

7.5% NaHCO₃ solution	`2.67`	ml
TPP/VFA mixture (see below)	`2.0`	ml
Rabbit serum heat-inactivated at 56C for 1 hr	`2.0`	ml

TPP/VFA mixture:

0.2% Thiamine pyrophosphate	`1.5`	ml
VFA solution (see below)	`1.0`	ml
Distilled water	`8.4`	ml

Mix together and filter-sterilize. Store at -20C.
VFA solution:

Isobutyric acid	`0.5`	ml
2-Methylbutyric acid	`0.5`	ml
Isovaleric acid	`0.5`	ml
Valeric acid	`0.5`	ml
0.1 N NaOH	`100.0`	ml

Add volatile fatty acids to about 70-80 ml of NaOH solution and mix. Add NaOH solution to bring final volume to 100 ml. If not used the same day, filter-sterilize and store at 4C.