|Pancreatic digest of casein||7 g|
|Yeast extract||3 g|
|Sodium chloride||5 g|
|Sodium deoxycholate||0.6 g|
|Crystal violet||0.002 g|
|Agar||12 to 18 g|
|Water, distilled||1000 mL|
Final pH is 7.0 ± 0.2 at 25°C. Agar amount can be adjusted depending on the gel strength. Different commercially available media may contain different amount of agar.
Grind the 5-bromo-4-chloro-3-indolyl-α-D-glucopyranoside with sodium chloride in a pestle and mortar to yield evenly distributed fine powder. Dissolve the other components in the water by boiling, remove from the heat and add the sodium chloride and 5-bromo-4-chloro-3-indolyl-α-D-glucopyranoside mixture. Adjust the pH, if necessary, so that it is 7.0 ± 0.2 at 25°C after sterilization. Autoclave at 121°C for 15 minutes. Cool the medium to between 44°C and 47°C. Mix well and pour about 15 mL to 20 mL into sterile Petri dishes and allow to solidify on a cool even surface.
After the plates have been poured and dried upside down in the dark at room temperature for up to 48 h, they can be placed in petri plate sleeves (cutting a 0.5" to 1" hole in the sleeves to allow condensation to escape) and stored upside down at 2-8 ºC in the dark for up to 2 weeks.
Enterobacter sakazakii Isolation Agar (ESIA) is available commercially from manufacturers such as Oxoid, Lenexa, KS (Cat. No. CM1134) and Millipore Sigma, Burlington, MA (Cronobacter spp. ChromoSelect Agar, Cat. No. 14703). Follow the manufacturer’s instructions if using commercial dehydrated medium. Media made from manufacturers following the recipe above can be used after verification.