How to prepare tryptic Soy Agar(TSA) : Principle,Composition and Colony response

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Tryptic soy agar MLTGEEKS

Tryptic Soy Agar plates abbreviated TSA is a general purpose growth medium suitable for the isolation of fastidious and non fastidious microorganisms,culture of both aerobes and anaerobes microorganism.Tryptic Soy Agar slant or Tryptic soy broth (tsb broth) is mostly used for the maintenance of stock culture such as cultivating,storing and shipment of bacterial cultures.This media is also recommended in the Manual of Clinical microbiology and diagnostic microbiology to be used in antibiotic susceptibility and sterility testing and can be used as a blood agar plate base by adding 7% sterile blood to the molten medium which has been cooled to a temperature of approximately 45° C .It can also be used in the detection of haemolysis with an addition of 0.7g lecithin and 5g of Polysorbate same as it can be used for the preparation of chocolate agar because it contains no added carbohydrate

Principle of Tryptic Soy Agar

  • The casein and soy peptones provide amino acids,nitrogen and peptides necessary for the growth of a wide variety of organisms.
  • Dextrose is a ready source of energy for the microorganism.
  • Sodium chloride supplies essential electrolytes and maintains osmotic equilibrium of the medium.
  • It may be supplemented with blood which facilitate the growth of more fastidious bacteria or antimicrobial agents to permit the selection of various microbial groups from mixed flora.

Composition of Tryptic Soy Agar

The following are tryptic soy agar ingredients used for it composition

Casein Peptone15.0 g
Sodium Chloride5.0 g
Soy Peptone5.0 g
Agar15.0 g
Demineralized Water1000.0 ml

How to Prepare Tryptic Soy Agar

  1. Start by suspending 45 grams in 1000 ml distilled water.
  2. Heat to boiling in order to completely dissolve the medium.
  3. Sterilize by autoclaving at 15 lbs pressure (121°C) for 15 minutes.
  4. Now cool to 45-50°C. Mix well and pour into sterile Petri dish with agar filled.
Also read  Bacteria motility test (Hanging drop technique)

Appearance of Medium after preparation

The growth medium appears light yellow coloured clear to slightly opalescent gel after preparation in petri dish or  tube

Tryptic Soy Agar 
Tryptic Soy broth 

How to Inoculate on Tryptic Soy Agar

  1. Inoculate and streak the specimen as soon as possible after it is received in the laboratory
  2. If the material is being cultured from a swab, you can roll the swab over a small area of the agar surface and then streak for isolation and spreading
  3. Incubate plates aerobically at temperature 33-37°C for 18-24 hours.
  4. Observe for typical colony morphology.
  5. For pour tube preparation,Melt the pour tube in a boiling water bath and cool to 45-50°C.
  6. Mix and dispense into a sterile petri dish and proceed with the instructions above.

Plate reading and Colonies morphology

MicroorganismIncubation timeGrowth Result
Bacillus subtilis Aerobic, 18-24 h 33-37°CGood growth
Aspergillus brasiliensis Aerobic, up to 72 h 25-30°CGood growth
Candida albicansAerobic, up to 72 h 25-30°CGood growth
Escherichia coli Aerobic, 18-24 h 33-37°C Good growth
Pseudomonas aeruginosaAerobic, 18-24 h 33-37°C Good growth
Staphylococcus aureusAerobic, 18-24 h 33-37°C Good growth
Escherichia coli on TSA
Escherichia coli on TSA
Pseudomonas aeruginosa on TSA
Pseudomonas aeruginosa on TSA


  • United States Pharmacopeia-National Formulary. 2007. 31st ed. United States Pharmacopeial Convention, Rockville, MD.
  • Murray, P.R., E.J. Baron, J.H. Jorgensen, M.L. Landry, and M.A. Pfaller. 2007. Manual of Clinical Microbiology. 9th ed. ASM Press, Washington, D.C.
  • Forbes, B.A., D.F. Sahm, and A.S. Weissfeld. 2007. Bailey and Scott’s Diagnostic Microbiology. 12th ed. Mosby Elsevier, St. Louis, MO
  • MacFaddin, J.F. 1985. Media for Isolation-Cultivation-Identification-Maintenance of Medical Bacteria. Vol. 1. Williams & Wilkins, Baltimore, MD.

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About the Author: Arthur Westmann

DEFFE ARTHUR (AMOEBAMANN) is the founder and author of MLTGEEKS and MLTEXPO.He’s from Cameroon and is currently a Final year State Medical Laboratory Technician (MLT MA). Beyond lab works, he’s a passionate internet user with a keen interest in web design and blogging. Furthermore He likes traveling, hanging around with friends and social networking to do in his spare time.

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