OXIDASE TEST

OXIDASE TEST

AIM

            To test the Oxidase enzyme producing efficiency of Bacteria.

PRINCIPLE    

            The Oxidase test is used to identify bacteria that produce Cytochrome - c oxidase, an enzyme of the bacterial Electron transport chain. In 1928, Gordon and McLeod introduced the use of a dimethyl-pphenylenediamine dihydrochloride solution to test for the presence of oxidase systems.  When the enzyme is present, the Cytochrome - c oxidase oxidizes the reagent Tetramethyl-p-phenylenediamine dihydrochloride to Indophenols (purple color end product). When the enzyme is not present, the reagent remains reduced and is colorless.

           Bacteria which contain Cytochrome - c as part of their respiratory chain are Oxidase positive and turn the reagent Blue/purple. Organisms lacking Cytochrome - c as part of their respiratory chain do not oxidize the reagent, leaving it colorless within the limits of the test, and are Oxidase negative. Examples for Oxidase positive bacteria are Pseudomonas sp., Vibrio sp., Neisseria gonorrhoeae, Campylobacter sp., Moraxella sp. and Pasteurella sp.

 MATERIALS REQUIRED

1. Test bacteria (24 hours culture only)
2. Oxidase disc (Tetramethyl-p-phenylenediamine dihydrochloride)
3. Glass slide
4. Sterile Wooden stick or Glass rod

PROCEDURE

a)   Pick the bacterial colony by using sterile wooden stick or glass rod and dip on the surface of Oxidase disc (Tetramethyl-p-phenylenediamine). Do not pick the inoculum with a metal loop because the Oxidase reagent may react with the metal.

b)     Observe inoculated area of Oxidase disc for a color change to deep blue or purple within 10 - 30 seconds.

OBSERVATION AND RESULTS

·       Oxidase Positive Test – Blue or purple colour with 10 – 30 seconds.

·       Oxidase Negative Test – No blue or purple colour.

Figure – 1: Oxidase test (Source: orbitbiotech.com)