# 689: ACE MEDIUM


## Main sol. 689

0.33	g	KCl	
6	g	MgCl2 x 6 H2O	
1	g	MgSO4 x 7 H2O	
0.25	g	NH4Cl	
0.14	g	CaCl2 x 2 H2O	
0.14	g	K2HPO4	
18	g	NaCl	
2	ml	Fe(NH4)2(SO4)2 x 6 H2O (0.1% w/v)	
1	g	Yeast extract (Difco)	
1	mg	Resazurin	
0.5	g	L-Cysteine HCl x H2O	
2	g	NaHCO3	
10	g	Glucose	
1	ml	**Wolin's vitamin solution (10x)**	
1000	ml	Distilled water	

1. Dissolve ingredients except cysteine, bicarbonate, glucose and vitamins, then sparge medium with 100% N2 gas to make it anoxic. Add cysteine and sodium bicarbonate, then adjust pH to 7.5. Dispense medium in anoxic Hungate-type tubes under 100% N2 gas and autoclave. A precipitate forms which will redissolve slowly over about 3 days. Before use inject separately sterile glucose solution (20% w/v) and vitamins prepared under 100% N2 gas atmosphere. Vitamins should be sterilized by filtration. Check medium pH and adjust pH to 7.6, if necessary.


## Wolin's vitamin solution (10x)

20	mg	Biotin	
20	mg	Folic acid	
100	mg	Pyridoxine hydrochloride	
50	mg	Thiamine HCl	
50	mg	Riboflavin	
50	mg	Nicotinic acid	
50	mg	Calcium D-(+)-pantothenate	
1	mg	Vitamin B12	
50	mg	p-Aminobenzoic acid	
50	mg	(DL)-alpha-Lipoic acid	
1000	ml	Distilled water