Mueller Hinton Agar : Composition, Preparation and Uses

Mueller Hinton Agar (MHA) is a common culture medium used for antimicrobial susceptibility testing (AST) of bacteria. Its composition provides optimal conditions for the growth of most clinically significant bacteria. In this article, we explore its principle, its preparation, its interpretation and its different uses.

◉ Overview

◉ Preparation and Composition of Mueller Hinton Agar

Suspend the components, dehydrated powder, in water (38 grams in 1000 ml of purified / distilled water). The medium is boiled for a few seconds until the ingredients are completely dissolved. Sterilize by autoclaving at 15 lbs (121 ° C) pressure for 15 minutes.

Cool to 47 ° C, mix well before pouring into sterile Petri dishes.

◈ A variety of supplements can be added to Mueller Hinton Agar , including 5% defibrinated sheep or horse blood, 1% growth supplement, and 2% sodium chloride.

◈ Starch is added to absorb all toxic metabolites produced

Mueller Hinton Agar's standardized composition and consistent performance make it an ideal medium for AST, enabling accurate and reliable interpretation of antimicrobial susceptibility results in clinical microbiology laboratories.

◉ Procedure

◈ A standardized suspension of the organism is buffered over the entire surface of the medium.

◈ Paper discs impregnated with specific amounts of antibiotics or other antimicrobial agents are placed on the surface backing.

◈ La boite est incubée et les zones d'inhibition autour de chaque disque sont mesurées. Détermination de la susceptibilité est faite en comparant la taille de zone obtenue aux tailles de zone dans CLSI, EUCAST


Antibiotic Susceptibility Testing

Limits of use

Some bacteria may not thrive on HD media due to their nutritional requirements. It is then recommended to use an appropriate medium: HTM for Haemophilus, Mueller-Hinton supplemented with 5% sheep blood for Streptococcus pneumoniae and ß hemolytic streptococci, GC agar for Neisseria gonorrhoae. These recommendations are well described in the various CLSI, CA-SFM or EUCAST standards.


  1. CLSI- Clinical & Laboratory Standards Institute Document M100
  2. The European Committee on Antimicrobial Susceptibility Testing - EUCAST- 2021
  3. Mueller, J. H. ; Hinton, Jane.- A Protein-Free Medium for Primary Isolation of the Gonoeoceus and Meningococcus.
  4. A. W. Bauer et al Antibiotic Susceptibility Testing by a Standardized Single Disk Method
  5. HUMEAU - Gélose Mueller-Hinton Fiche technique
  6. MILLIPORE - Mueller-Hinton AGAR
  7. northeast laboratory services - Mueller-Hinton AGAR
  8. Neogen- Mueller Hinton Agar II