Catalase and Oxidase Biochemical Tests

Catalase and oxidase tests are common biochemical tests used in microbiology to identify and differentiate between different bacterial species based on their metabolic characteristics.

Here's a basic protocol for performing the catalase and oxidase tests:

Catalase Test:

​

The catalase test is used to determine if bacteria produce the enzyme catalase, which breaks down hydrogen peroxide into water and oxygen.

Materials Needed:

Bacterial culture (overnight culture or fresh culture)
Sterile inoculating loop or bacteriological loop
Hydrogen peroxide solution (3% w/v)
Microscope slide or small test tube

Procedure:

Preparation:
a. Prepare a fresh bacterial culture or use an overnight culture.
b. If using an overnight culture, ensure that it is still in the exponential growth phase.

Testing:
a. Gently emulsify a small amount of bacterial culture using a sterile loop.
b. Place a drop of the bacterial culture on a microscope slide or in a small test tube.
c. Add a drop of 3% hydrogen peroxide to the bacterial culture. Observe for rapid bubbling or effervescence.

Interpretation:

If bubbling occurs immediately upon adding hydrogen peroxide, the bacterium is catalase-positive, indicating the presence of the catalase enzyme.
If no bubbling occurs, the bacterium is catalase-negative.

Oxidase Test:
The oxidase test is used to identify bacteria that possess the enzyme cytochrome c oxidase, which is involved in the electron transport chain.

Materials Needed:

Oxidase reagent (tetramethyl-p-phenylenediamine dihydrochloride)
Sterile swabs or inoculating loops
Bacterial culture (overnight culture or fresh culture)

Procedure:

Preparation:
a. Prepare a fresh bacterial culture or use an overnight culture.
b. If using an overnight culture, ensure that it is still in the exponential growth phase.

Testing:
a. Using a sterile swab or inoculating loop, transfer a small amount of bacterial culture onto a piece of filter paper or directly onto the oxidase test strip.
b. Add a drop of oxidase reagent onto the bacterial culture on the filter paper or test strip. Observe for color change.

Interpretation:

If the bacterial culture turns dark blue or purple within 20-30 seconds, the bacterium is oxidase-positive, indicating the presence of cytochrome c oxidase.

If there is no color change, or only a faint color change occurs, the bacterium is oxidase-negative.
Always follow proper laboratory safety guidelines and sterile techniques while working with bacterial cultures and reagents.

Interpretation of results should be done in the context of other test results and clinical information. The protocol may need adjustments based on specific bacterial strains and testing conditions.