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Important: Acidity inhibits oxidase enzyme activity. The oxidase test must not be performed, therefore, on colonies that produce fermentation on carbohydrate – containing media, such as sucrose fermenting V.cholerae colonies on TCBS medium, Subinoculation on nutrient agar is required before the oxidase test can be performed reliably. Non – fermenting colonies, however, can be tested. Colonies tested from a medium that contains nitrate may give unreliable oxidase test results.

Required

Oxidase reagent

Freshly prepared

This is a 10g/l dihydrochloride.

solution of tetramethyl –p-phenylenediamine

Note: Oxidase reagent is easily oxidized. When oxidized, it is blue in colour and must not be used.

Method

Place a piece of filter paper in a clean petri dish and add 2 or 3 drops of freshly prepared oxidase reagent. Using a piece of stick or glass rod (not an oxidized wire loop), remove a colony of the test organism, and smear it on the filter paper. Look for the development of a blue – purple colour within a few seconds.

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