Microbiology Testing FAQs — Methods & Media | HiMedia USA

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Why shouldn't EMB Agar be autoclaved?

EMB Agar should NOT be autoclaved because: (1) Excessive heat precipitates eosin Y and methylene blue dyes, (2) Overheating reduces selectivity, (3) Can inhibit organism growth, (4) Color reactions may be obscured. Proper preparation: Heat to dissolve completely (boiling), cool immediately to 45-50°C, pour plates. Brief boiling is sufficient for sterilization. Autoclaving EMB results in: dark precipitate in medium, poor selectivity, false-negative metallic sheen. This is why many labs prefer RTU EMB plates - eliminates preparation errors.
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How do I prepare MacConkey Agar from powder?

Suspend 50g of MacConkey Agar powder in 1000mL purified water. Heat to boiling to dissolve completely (do NOT autoclave). Cool to 45-50°C. Mix well and pour into sterile petri dishes (approximately 20mL per 90mm plate). Final pH 7.1±0.2. Medium should be pinkish-red (from neutral red indicator). Do not overheat or autoclave MacConkey - excessive heating destroys bile salts and crystal violet reducing selectivity. Once poured and solidified, store plates inverted at 2-8°C. Use within 1-2 weeks or as validated.
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