The oxidase test uses a colorless reagent tetra-methyl- p-phenylenediamine dihydrochloride (TMPD), which acts as an artificial donor for cytochrome c. When cytochrome c oxidase is present, it oxidizes TMPD, changing it from colorless to a purple or dark blue compound. In the absence of the enzyme, TMPD remains reduced and colorless.
Hence, if a blue or purple color develops on adding TMPF, it indicates that the bacteria possess the cytochrome c oxidase enzyme. If there’s no color change, it indicates that the bacteria lacks this enzyme.