Cat. no. C6420 CRITERION™ Mueller Hinton Agar 76gm
Cat. no. C6421 CRITERION™ Mueller Hinton Agar 500gm
Cat. no. C6422 CRITERION™ Mueller Hinton Agar 2kg
Cat. no. C6423 CRITERION™ Mueller Hinton Agar 10kg
Cat. no. C6424 CRITERION™ Mueller Hinton Agar 50kg


Hardy Diagnostics CRITERION™ Mueller Hinton Agar is recommended for use in the cultivation of a wide variety of microorganisms. Mueller Hinton Agar is recommended for disk diffusion sensitivity testing of non-fastidious organisms.

This dehydrated culture medium is a raw material intended to be used in the making of prepared media products, which will require further processing, additional ingredients, or supplements.


Mueller and Hinton developed Mueller Hinton Agar in 1941 to be a protein-free medium for isolating pathogenic strains of Neisseria.(10) In recent times this media has been used in standardized antimicrobial disk susceptibility testing, as described by Bauer, Kirby, et al.(5) Barry and Fay investigated the effects of altering the depth of plated Mueller Hinton Agar on disk diffusion testing, and determined a standardized depth of approximately four millimeters to be sufficient.(4) In 1970 Dewees, et al., studied the effect of storage on Mueller Hinton Agar plates used for antimicrobial disk diffusion zone sizes. Their findings indicated commercially manufactured Mueller Hinton Agar plates were suitable for use in routine susceptibility testing.(6) In addition to the above criteria, CRITERION™ Mueller Hinton Agar meets the standards of performance established by the Clinical Laboratory Standards Institute (CLSI), documents M2, M100, and ISO 16782.(11,15,16)

CRITERION™ Mueller Hinton Media contains beef infusion and casamino acids, and starch. Starch acts as a colloid that protects against toxic material in the medium. Beef infusion and casamino acids are provided as a source of energy and nutrients. Agar is added as a solidifying agent. The levels of tetracycline and sulfonamide inhibitors, thymidine, thymine, magnesium and calcium ions are controlled so as not to interfere with susceptibility testing and to yield good growth.

The Kirby-Bauer antimicrobial disk diffusion procedure is used with Mueller Hinton Agar plates. Disk diffusion testing is based on an antimicrobial diffusing through an agar gel, when placed on the agar surface after it has been impregnated onto a filter paper disk.(11,14) Zone diameters established for each antimicrobial determining resistant, intermediate, and sensitive results for pathogenic microorganisms are listed in the Clinical Laboratory Standards Institute (CLSI), document M100.(15)


Gram weight per liter: 38.0gm/L
Acid Hydrolysate of Casein 17.5gm
Beef Extract 2.0gm
Starch 1.5gm
Agar 17.0gm

Final pH 7.3 +/- 0.1 at 25ºC.

* Adjusted and/or supplemented as required to meet performance criteria.


Store the sealed bottle(s) containing culture medium at 2-30ºC. Dehydrated culture medium is very hygroscopic. Keep lid tightly sealed. Protect dehydrated culture media from moisture and light. The dehydrated culture media should be discarded if it is not free-flowing or if the color has changed from its original beige.

Store the prepared plated media at 2-8ºC.



1. Suspend 38.0gm of the dehydrated culture media in 1 liter of distilled or deionized water.

2. Heat to boiling and mix to dissolve completely.

3. Sterilize in the autoclave at 121ºC. for 15 minutes. Avoid overheating.

4. Cool to 50-55ºC. and dispense to give an approximate depth of 4mm. Refer to CLSI (NCCLS) document M2-A.(11)


For information on procedures and interpretation of results, consult listed references or refer to the prepared media Instructions for Use (IFU) for Cat. No. H11.


The disk diffusion method should not be used for obligatory anaerobes, slow growing organisms, and capnophiles. This method was standardized for facultative organisms or rapid growing aerobes.

When using the disk diffusion method, technical human errors may compromise reliability and accuracy. The following errors are common sources encountered in the clinical microbiology laboratory, and must be watched for: improper disk storage, inoculum not properly adjusted (too light or too heavy), incubation temperature deviating from 35-37ºC., use of an increased CO2 atmosphere, reading plates before or after the full 16-18 hours of incubation, transcribing errors, reader error when measuring zone diameters, deterioration of the McFarland Turbidity Standard, and contamination or mutation in the control strain(s).


Standard microbiological supplies and equipment such as autoclaves, incinerators, and incubators, etc., are not provided.


Test Organisms Inoculation Method* Incubation Results
Time Temperature Atmosphere
Escherichia coli
ATCC® 25922
F 24hr 35°C Aerobic Growth**
Escherichia coli
ATCC® 35218
F 24hr 35°C Aerobic Growth**
Staphylococcus aureus
ATCC® 25923
F 24hr 35°C Aerobic Growth**
Pseudomonas aeruginosa
ATCC® 27853
F 24hr 35°C Aerobic Growth**
Enterococcus faecalis
ATCC® 29212
F 24hr 35°C Aerobic Growth**
Enterococcus faecalis
ATCC® 33186
F 24hr 35°C Aerobic Growth**
Staphylococcus aureus
ATCC® NCTC 12493
F 24hr 35°C Aerobic Growth**
Staphylococcus aureus
ATCC® 43300
F 24hr 35°C Aerobic Growth**

**For appropriate disk diffusion ranges, consult CLSI M100 and ISO 16782.(15,16)

User Quality Control


CRITERION™ Mueller Hinton Agar powder should appear homogeneous, free-flowing and beige, with a few dark specks, in color. The prepared media should appear translucent, and light amber in color.


1. Anderson, N.L., et al. Cumitech 3B; Quality Systems in the Clinical Microbiology Laboratory, Coordinating ed., A.S. Weissfeld. American Society for Microbiology, Washington, D.C.

2. Jorgensen., et al. Manual of Clinical Microbiology, American Society for Microbiology, Washington, D.C.

3. Tille, P., et al. Bailey and Scott's Diagnostic Microbiology, C.V. Mosby Company, St. Louis, MO.

4. Barry and Fay. 1973. Am. J. Clin. Pathol.; 50:196.

5. Bauer, A.W., W.M.M. Kirby, et al. 1966. Am. J. Clin. Pathol.; 45:493-496.

6. Dewees, et al. 1970. Effect of storage of Mueller Hinton Agar plates on zone sizes for antimicrobial testing. Appl. Microbiol.; 30:203.

7. Isenberg, H.D. Clinical Microbiology Procedures Handbook, Vol. I, II & III. American Society for Microbiology, Washington, D.C.

8. Koneman, E.W., et al. Color Atlas and Textbook of Diagnostic Microbiology, J.B. Lippincott Company, Philadelphia, PA.

9. Methods for Dilution Antimicrobial Susceptibility Test For Bacteria That Grow Aerobically, M7-current edition. Clinical Laboratory Standards Institute (CLSI - formerly NCCLS), Villanova, PA.

10. Mueller, J.H. and J. Hinton. 1941. A protein-free medium for primary isolation of the Gonococcus and Meningococcus. Proc. Soc. Exp. Diol. and Med.; 48:330-333.

11. Performance Standards for Antimicrobial Disk Susceptibility Tests, 7th ed. M2-A7. 2000. Clinical Laboratory Standards Institute (CLSI - formerly NCCLS), Wayne, PA.

12. Quality Assurance for Commercially Prepared Microbiological Culture Media, M22. Clinical and Laboratory Standards Institute (CLSI - formerly NCCLS), Wayne, PA.

13. Ryan, K.J., et al. 1970. Disk sensitivity testing. Hosp. Prac.; 5:91-100.

14. Standard Disk Susceptibility Test, The Federal Register, September 30, 1972; 37(191):20527-20529.

15. Performance Standards for Antimicrobial Susceptibility Testing. M100. Clinical and Laboratory Standards Institute (CLSI), Wayne, PA.

16. ISO 16782. Clinical laboratory testing - Criteria for acceptable lots of dehydrated Mueller-Hinton agar and broth for antimicrobial susceptibility testing.

ATCC is a registered trademark of the American Type Culture Collection.