Catalog Number: 1007317
Mueller-Hinton agar

For use in the tests for organisms susceptibility to antimicrobial agents by the disk diffusion method.1 Also for the cultivation of Neisseria.

Mueller-Hinton Agar is a medium very rich in nutrients that was originally recommended for the isolation and development of gonococci and meningococci. It is used primarily for sensitivity testing of microorganisms.

Approximate formula g/l:
    Beef Infusion
3.0Casein Peptone H17.5
Final pH 7.4 + 0.2


Suspend 38 g of the medium in one liter of distilled or deionized water. Mix well and heat with frequent agitation. Boil for one minute and sterilize at 121o C (15 lbs. of pressure) for 15 minutes.

Cool to 40-45o C and pour into Petri dishes. If desired, plates or tubes of "chocolate" agar can be prepared by heating the medium containing sheep blood in a waterbath at 80o C for 10 minutes. Do not overheat the medium.


Mueller-Hinton Agar can be used to cultivate Nesseria specimens. It is recommended to incubate the plates at 35oC in a CO2 atmosphere. The medium should be maintained in a humid condition. This is the best attained by adding a damp sponge or cotton cloth on the bottom of the candle jar or other appropriate containers.

For susceptibility testing follow the NCCLS (National Committee for Clinical Laboratory Standards) guidelines, Performance Standards for Antimicrobial Disk Tests, M2-T4. Plates should be examined after 16 to 18 hours of incubation. After this time the zones of inhibition must be checked because the microorganism can begin to grow when the concentration of the antimicrobial agent decreases.

Better results for isolating pathogenic neisserias are obtained by preparing a chocolate agar of Mueller-Hinton adding to every 100 ml of final medium 1.0 ml of Polienrichment + 1.0 ml of VCN suspension.


The use of a suitable medium for testing the susceptibility of microorganisms to sulfonamides and trimethoprim is essential. Antagonism to sulfonamide activity is demonstrated by para-aminobenzoic acid (PABA) and its analogs. Reduced activity of trimethoprim, resulting in smaller inhibition zones and innerzonal colonies, is demonstrated on unsuitable Mueller Hinton medium possessing high levels of thymidine. Both the PABA and thymine/thymidine content in Mueller Hinton Agar are reduced to a minimum, thus markedly reducing the inactivation of sulfonamides and trimethoprim when the media is used for testing the susceptibility of bacterial isolates to these antimicrobics.


The following are typical results obtained in the performance of the medium from type cultures after incubation at a temperature of 37oC and observed after 24 hours.

Escherichia coli ATCC 25922
good to excellent
Neisseria meningitidis ATCC 13090
good to excellent
Staphylococcus aureus ATCC 25923
good to excellent
Streptococcus faecalis ATCC 33186
good to excellent
Streptococcus pneumoniae ATCC 6303
good to excellent
Pseudomonas aeruginosa ATCC 27853
good to excellent

  1. NCCLS Approved Standard: ASM-2, Performance standards for antimicrobic disc susceptibility tests, 2nd Ed. Villanova, PA., National Committee for Clinical Laboratory Standards (1979).
  2. Tentative Standard, Methods for dilution antimicrobial susceptibility tests for bacteria that grow aerobically. Villanova, PA., National Committee for Clinical Laboratory Standards (1983).
  3. Gordon and Hine, Brit. Med. J., v. 678 (1916).
  4. Goodale, Gould, Schwab, and Winter, JAMA, v. 123: 547 (1943).
  5. Bauer, et. al., Am. J. Clin. Path., v. 45: 493 (1966).
  6. Mueller and Hinton, "A Protein-Free Medium for Primary Isolation of the Gonococcus and Meningococcus." Proc. Soc. Exp. Biol. and Med., v. 48:330 (1941).
  7. Harris and Coleman, Diagnostic Procedures and Reagents, 4th Edition, APH, Inc. New York, 1963.
  8. National Committee for Clinical Laboratory Standards. M2-4T. Vol. 8. No. 7


Catalog NumberDescriptionSize
1007317Mueller Hinton Agar500 g
1007417Mueller Hinton Broth500 g