Mueller Hinton Agar (MHA)

Mueller Hinton Agar (MHA) Powder

Standard Reference Medium for Antimicrobial Susceptibility Testing

Catalog Number: AS-1299
HS / AHECC Code: 3821.00.00
Form: Dehydrated Powder
Grade: Microbiology / AST Reference Medium

Product Overview

Mueller Hinton Agar (MHA) is the internationally accepted standard solid medium for antimicrobial susceptibility testing (AST) of non-fastidious bacteria. It is the reference agar used for the Kirby–Bauer disk diffusion method, MIC determination, and gradient diffusion techniques.

Mueller Hinton Agar is specified as the official testing medium in major global standards including CLSI M02 / M07, EUCAST, ISO 20776-1, and is recommended by regulatory bodies such as the FDA and WHO for standardized antibiotic susceptibility testing.

Due to its carefully controlled composition, low thymidine content, and regulated cation levels, Mueller Hinton Agar ensures reliable antibiotic diffusion and reproducible inhibition zones, making it the benchmark medium used in clinical microbiology laboratories, research institutions, pharmaceutical quality control, and antimicrobial resistance studies worldwide.

Key Features & Benefits

• International reference medium for antimicrobial susceptibility testing
• Standardized composition compliant with CLSI, EUCAST, and ISO guidelines
• Low thymidine and thymine content for accurate sulfonamide and trimethoprim testing
• Controlled calcium and magnesium concentrations for reproducible aminoglycoside and tetracycline activity
• Starch component absorbs bacterial toxins and improves inhibition zone clarity
• Excellent antibiotic diffusion properties for Kirby–Bauer disk testing
• Reliable batch-to-batch performance for laboratory standardization

Typical Applications

Antimicrobial Susceptibility Testing

Mueller Hinton Agar is widely used for:

• Kirby–Bauer disk diffusion antibiotic testing
• Minimum Inhibitory Concentration (MIC) testing
• Gradient diffusion strip testing (Etest)
• Antimicrobial resistance surveillance

Routine Clinical Microbiology

Testing susceptibility of:

• Staphylococcus aureus
• Escherichia coli
• Pseudomonas aeruginosa
• Acinetobacter spp.
• Enterococcus spp.
• Enterobacteriaceae family organisms

Specialized Testing Variants

Modified Mueller Hinton Agar formulations are used for specific organisms:

• MHA + 5% defibrinated blood → Streptococcus spp.
• MH-F medium (Mueller Hinton + supplements) → Haemophilus spp.
• MHA + 2% NaCl → MRSA screening (oxacillin / methicillin resistance)

Principle of the Medium

Mueller Hinton Agar provides a nutritionally adequate yet controlled environment that supports bacterial growth while allowing uniform antibiotic diffusion through the agar matrix.

The medium contains acid hydrolysate of casein and beef infusion which provide amino acids and nutrients required for microbial growth. Starch absorbs bacterial toxins that may interfere with antibiotic activity, resulting in clear and well-defined inhibition zones.

Controlled concentrations of divalent cations such as calcium and magnesium are critical for reproducible susceptibility testing, particularly for aminoglycosides, polymyxins, and tetracycline antibiotics.

Low thymidine and thymine levels prevent false resistance results when testing folate pathway inhibitors such as sulfonamides and trimethoprim.

Typical Composition (per litre)

Final pH: 7.3 ± 0.1 at 25 °C

Cation Content (after sterilization)

These controlled levels ensure reproducible antibiotic activity in susceptibility testing.

Method of Preparation

1. Suspend 38.0 g of Mueller Hinton Agar powder in 1 litre of purified water.
2. Heat with frequent agitation and boil for 1 minute to dissolve completely.
3. Sterilize by autoclaving at 121 °C for 15 minutes.
4. Cool the medium to 45–50 °C.
5. Pour 25–30 mL into 90–100 mm Petri dishes to obtain a final agar depth of 4.0 ± 0.5 mm (critical for disk diffusion testing).
6. Allow plates to solidify and dry before use.

Incubation Conditions

Temperature: 35 ± 1 °C
Atmosphere: Ambient air (unless otherwise specified)
Typical incubation time: 16–20 hours

Storage & Stability

Dehydrated medium:
Store tightly closed at 15–30 °C in a dry environment.

Prepared agar plates:
Store at 2–8 °C in sealed plastic bags.
Prepared plates are typically suitable for 7 days under routine laboratory conditions.

Intended Use

For laboratory antimicrobial susceptibility testing only.

Not intended for human, veterinary, pharmaceutical, food, or therapeutic applications.

Quality & Compliance

Manufactured under controlled laboratory conditions to ensure reproducible performance and consistent quality.

Meets performance requirements specified by:

• CLSI M02 / M07
• EUCAST standards
• ISO 20776-1 guidelines

Performance comparable to leading international microbiology media manufacturers.