TRYPTOSE PHOSPHATE BROTH

Code: CM0283

A buffered glucose broth for the cultivation of fastidious bacteria.

* Adjusted as required to meet performance standards

Directions
Dissolve 29.5g in 1 litre of warm distilled water and distribute into final containers. Sterilise by autoclaving at 121°C for 15 minutes.

For the cultivation of anaerobes, if the reconstituted medium has been stored prior to use, remove dissolved oxygen by placing the tubes in a boiling water bath for 15 minutes and cool without agitation before inoculation.

Description
A buffered dextrose broth for use as an adjuvant to tissue culture media and for the cultivation of fastidious bacteria. Ginsberg et al.¹ maintained tissue cultures of HeLa cells for at least 10 days in a mixture of Tryptose Phosphate Broth 15- 25%, Scheren maintenance solution 67.5-77.5% and chicken serum 7.5%. The cells increased 3-5 fold in number during this period, smaller quantities of ARD, AD and type 1 poliomyelitis virus could be detected and more ARD virus could be propagated in HeLa cells in the Tryptose Phosphate.

Broth supplemented medium
Tryptose Phosphate Broth is recommended for the cultivation of streptococci, pneumococci, meningococci and other fastidious organisms. Tryptose Phosphate Broth with added agar and sodium azide is recommended for the isolation of pathogenic streptococci from cheese and other dairy products^2,3.

Tryptose Phosphate Broth with added agar is also recommended by the American Public Health Association for the examination of throat swabs and blood for Streptococcus pneumoniae and as a growth medium for pneumococci prior to the bile solubility test.

Tryptose Phosphate Agar Broth may also be employed for the emulsification of cheese prior to the plate isolation method for Brucella species².
The small proportion of agar (0.1-0.2%) necessary for the above methods may be most conveniently added to the Oxoid broth before sterilisation, in the form of Oxoid Agar Tablets CM0049 (one tablet per 100 ml).

Technique
Examination of throat swabs for Streptococcus pneumoniae⁴
1. Prepare Tryptose Phosphate Broth in the usual manner but add a small amount of agar by dissolving 10 Agar Tablets CM0049 per litre of broth before autoclaving.
2. Place a pharyngeal or sputum swab in 3ml of Tryptose Phosphate (Agar) Broth and incubate for 2 hours at 35°C. If sufficient pneumococci are present, type directly or preferably employ the culture for mouse inoculation.

Examination of cheese for Lancefield Group A streptococci²
1. Heat 25ml of Tryptose Phosphate (Agar) Broth (prepared as above) to 45°C.
2. Immediately emulsify 5 grams of cheese in the broth, using a heavy sterile glass rod. Plate suitable dilutions of the emulsion on Tryptose Agar plates containing 1.5% agar and 0.04% of sodium azide.
Alternatively, add sterile aqueous sodium azide to the emulsion to give a final concentration of 2.5%. Incubate for 12-14 hours at 35°C, then shake and plate on Tryptose Agar plates containing 1.5% agar and 0.04% of sodium azide.
The sodium azide suppresses the growth of most bacteria except streptococci and some lactobacilli.

Blood Culture
1. Add up to 10ml of blood to 150ml of Tryptose Phosphate Broth in a 300ml flask or bottle.
2. Incubate and sub-culture on to other media in the usual manner, according to the exact purpose of the investigation.

Storage conditions and Shelf life
Store the dehydrated medium at 10-30°C and use before the expiry date on the label.
Store the prepared medium at 2-8°C.

Appearance
Dehydrated medium: Straw coloured, free-flowing powder
Prepared medium: Light straw coloured solution

Quality control

* This organism is available as a Culti-Loop®

References
1. Ginsberg H. S. et al. (1955) Proc. Soc. Exper. Biol. Med. 89. 66-71.
2. American Public Health Association (1953) `Standard Methods for the Examination of Dairy Products’ 10th ed., APHA Inc.,
New York, pp. 179, 180, 181.
3. Newman R. W. (1950) J. Milk Food Tech. 13. 226-233.
4. American Public Health Association (1953) `Diagnostic Procedures and Reagents’ 4th ed., APHA Inc., New York, p. 141.