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山東拓普生物工程有限公司

Shandong Tuopu Biol-Engineering Co.,Ltd

Anaerobiospirillum thomasii Medium
(DSMZ Medium 800)
Composition per 1070mL:
Pancreatic digest of casein..........................................................10.0g
Gelatin peptone...........................................................................10.0g
NaCl..............................................................................................5.0g
Yeast extract..................................................................................5.0g
Glucose .........................................................................................1.0g
L- Arginine .....................................................................................1.0g
Sodium pyruvate...........................................................................1.0g
Hemin.........................................................................................5.0mg
Menadione .................................................................................0.5mg
Fildes enrichment solution.....................................................100.0mL
NaHCO 3  solution .....................................................................50.0mL
Na2S·9H2O solution.................................................................10.0mL
Cysteine solution......................................................................10.0mL
pH 6.9 ± 0.2 at 25°C
Fildes Enrichment Solution:
Composition per 206.0mL:
Pepsin............................................................................................1.0g
NaCl (0.85% solution)...........................................................150.0mL
Sheep blood, defibrinated........................................................50.0mL
HCl.............................................................................................6.0mL
Source:  Fildes enrichment solution is available as a premixed powder
from BD Diagnostic Systems and Oxoid Unipath.
Preparation of Fildes Enrichment Solution: Combine compo-
nents. Mix thoroughly. Incubate at 56°C for 4 hr. Bring pH to 7.0 with
20% NaOH. Adjust pH to 7.2 with HCl. Do not autoclave. Add 0.25
mL of chloroform and store at 4°C. Before use, heat to 56°C to remove
chloroform.
Na 2 S·9H 2 O Solution :
Composition per 10.0mL:
Na 2 S·9H 2 O....................................................................................0.3g
Preparation of Na 2 S·9H 2 O Solution: Add Na 2 S·9H 2 O to dis-
tilled/deionized water and bring volume to 10.0mL. Mix thoroughly.
Autoclave under 100% N 2 for 15 min at 15 psi pressure–121°C. Cool
to room temperature.
Cysteine Solution:
Composition per 10.0mL:
L -Cysteine·HCl·H 2 O.....................................................................0.3g
Preparation of Cysteine Solution:  Add  L -cysteine·HCl·H 2 O to
distilled/deionized water and bring volume to 10.0mL. Mix thorough-
ly. Sparge with 100% N 2 . Autoclave for 15 min at 15 psi pressure–
121°C. Cool to room temperature.
NaHCO 3 Solution:
Composition per 100.0mL:
NaHCO 3 ........................................................................................5.0g
Preparation of NaHCO 3 Solution:  Add NaHCO 3 to distilled/de-
ionized water and bring volume to 100.0mL. Mix thoroughly. Sparge
with 80% N 2 + 20% CO 2 . Filter sterilize.
Preparation of Medium: Prepare and dispense medium under 80%
N 2 + 20% CO 2 gas atmosphere. Add components, except Fildes enrich-
ment solution, cysteine solution, NaHCO 3  solution, and Na 2 S·9H 2 O
solution, to distilled/deionized water and bring volume to 1.0L. Mix
thoroughly. Distribute into tubes or flasks. Autoclave for 15 min at 15
psi pressure–121°C. Cool while sparging with 80% N 2 + 20% CO 2 .
Aseptically and anaerobically add 100.0mL Fildes enrichment solution,
10.0mL cysteine solution, 50.0mL NaHCO 3  solution, and 10.0mL
Na 2 S·9H 2 O solution. Aseptically and anaerobically distribute to sterile
tubes or bottles.
Use: For the cultivation of Anaerobiospirillum thomasii.