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Standard Fluid Medium 10B (Shepard’s M10 Medium)

[所属分类:培养基配方] [发布时间:2021-8-25] [发布人:网站管理员2] [阅读次数:] [返回]

Standard Fluid Medium 10B

(Shepard’s M10 Medium)

山东拓普生物工程有限公司培养基配方 http://www.topbiol.com

Composition per 102.5mL:

Base solution............................................................................70.0mL

Horse serum, unheated.............................................................20.0mL

Fresh yeast extract solution......................................................10.0mL

Penicillin solution ......................................................................1.0mL

CVA enrichment.........................................................................0.5mL

L-Cysteine·HCl·H2O solution.....................................................0.5mL

Urea solution..............................................................................0.4mL

Phenol Red solution ...................................................................0.1mL

pH 6.0 ± 0.2 at 25°C

Base Solution:

Composition per 70.0mL:

Beef heart, solids from infusion.................................................... 5.0g

Peptone.......................................................................................... 1.0g

NaCl .............................................................................................. 0.5g

Preparation of Base Solution: Add components to distilled/deion

ized water and bring volume to 70.0mL. Mix thoroughly. Adjust pH to

5.5 with 2N HCl. Autoclave for 15 min at 15 psi pressure–121°C. Cool

to 45°–50°C.

Fresh Yeast Extract Solution:

Composition per 100.0mL:

Baker’s yeast, live, pressed, starch-free,..................................... 25.0g

Preparation of Fresh Yeast Extract Solution: Add the live Bak

er’s yeast to 100.0mL of distilled/deionized water. Autoclave for 90

min at 15 psi pressure–121°C. Allow to stand. Remove supernatant so

lution. Adjust pH to 6.6–6.8.

Penicillin Solution:

Composition per 10.0mL:

Penicillin G ....................................................................... 1,000,000U

Preparation of Penicillin Solution: Add penicillin to distilled/de

ionized water and bring volume to 10.0mL. Mix thoroughly. Filter ster

ilize.

CVA Enrichment:

Composition per liter:

Glucose ..................................................................................... 100.0g

L-Cysteine·HCl·H2O.................................................................... 25.9g

L-Glutamine................................................................................. 10.0g

L-Cystine·2HCl.............................................................................. 1.0g

Adenine......................................................................................... 1.0g

Nicotinamide adenine dinucleotide ............................................ 0.25g

Cocarboxylase............................................................................... 0.1g

Guanine·HCl ............................................................................... 0.03g

Fe(NO3)3 ..................................................................................... 0.02g

p-Aminobenzoic acid................................................................ 0.013g

Vitamin B12 ................................................................................. 0.01g

Thiamine·HCl ............................................................................3.0mg

Preparation of CVA Enrichment: Add components to distilled/

deionized water and bring volume to 1.0L. Mix thoroughly. Filter ster

ilize.

L-Cysteine·HCl·H2O Solution:

Composition per 10.0mL:

L-Cysteine·HCl·H2O ..................................................................... 0.2g

Preparation of L-Cysteine·HCl·H2O Solution: Add L-cyste

ine·HCl·H2O to distilled/deionized water and bring volume to 10.0mL.

Mix thoroughly. Filter sterilize.

Urea Solution:

Composition per 10.0mL:

Urea............................................................................................... 1.0g

Preparation of Urea Solution: Add urea to distilled/deionized wa

ter and bring volume to 10.0mL. Mix thoroughly. Filter sterilize.

Phenol Red Solution:

Composition per 10.0mL:

Phenol Red.................................................................................... 0.1g

Preparation of Phenol Red Solution: Add Phenol Red to dis

tilled/deionized water and bring volume to 10.0mL. Mix thoroughly.

Autoclave for 15 min at 15 psi pressure–121°C.

Preparation of Medium: To 70.0mL of cooled, sterile base solu

tion, aseptically add 20.0mL of sterile horse serum, 10.0mL of sterile

fresh yeast extract solution, 1.0mL of sterile penicillin solution, 0.5mL

of sterile CVA enrichment, 0.5mL of sterile L-cysteine·HCl·H2O solu

tion, 0.4mL of sterile urea solution, and 0.1mL of sterile Phenol Red

solution. Mix thoroughly. Aseptically distribute into sterile tubes or

flasks.

Use: For the isolation and cultivation of Ureaplasma urealyticum from

clinical specimens.

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Standard Fluid Medium 10B (Shepard’s M10 Medium)