 The cytochromes are iron-containing hemoproteins

that act as the last link in the chain of aerobic

respiration by transferring electrons (hydrogen) to
oxygen, with the formation of water.
 Cytochrome oxidase or Warburge’s respiratory
enzyme – the enzyme which reduces
molecular oxygen to water to compete the
last link in the chain of aerobic respiration
 The test is most helpful in screening colonies
suspected of being one of the Enterobacteriaceae
(all negative) and in identifying colonies suspected
of being a pseudomonas specie or a neisseria
specie which are all positive. (Pseudomonas, Vibrio,
Aeromonas, Neisseria and Comamonas)
 The cytochrome oxidase test utilize certain
reagent dyes, such as the “redux dye:
tetramethyl-paraphenylenediamine
dihydrochloride”, that substitute for oxygen as
artificial electron acceptors.

 In the reduced state the dye is colorless, however,

in the presence of cytochrome oxidase and
atmospheric oxygen, tetramethyl-
paraphenylenediamine dihydrochloride is
oxidized, forming indophenol blue color.
 Materials:

Culture plate of organism

Cytochrome oxidase paper strip (Pathotec oxidase
strip) (tetra-methyl-para-phenylene diamine
dihydrochloride)
 Procedures:

 Thetest is commonly performed by one of the

two methods:
 The direct plate technique in which 2 to 3
drops reagent are directly poured into the
isolated bacteria colonies growing on the plate
medium.
 The indirect paper strip procedure. In which a
few drops of the reagent are either added to a
filter paper trip or commercial disk or strips
impregnated which dried reagent are used.
 Ineither method, a loopful of suspected colony
is smeared into the reagent zone of the filter
paper.
Observations:

 Bacterial colonies having cytochrome oxidase

actively will develop a deep blue color to purple
to black at the inoculation site within seconds.