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SOP

Oxidase Test for Bacterial Identification

SOP for detecting cytochrome c oxidase to differentiate bacterial groups (e.g., Pseudomonas vs. Enterobacteriaceae).

Last verified 1 month ago 1 min read Microbiology Bacteriology Biochemical Testing #microbiology #Biochemical Test #oxidase test #pseudomonas
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Document Control
Version
1.0
Effective
Aug 28, 2026
Reviewed
Aug 29, 2026
Department
Microbiology
Principle

Cytochrome c oxidase oxidizes the reagent (tetramethyl-p-phenylenediamine) to form a dark purple indophenol compound.

Standard Operating Procedure: Oxidase Test

1. Purpose: Detect cytochrome c oxidase enzyme.

2. Specimen: Fresh 18-24h bacterial colony (avoid sugar-containing media like MacConkey).

3. Reagents: 1% Tetramethyl-p-phenylenediamine dihydrochloride (Kovacs reagent).

4. Procedure:

  • Place filter paper on a clean slide.
  • Add 1-2 drops of oxidase reagent to the paper.
  • Using a wooden stick or platinum loop, rub a heavy inoculum of the colony onto the wet paper.

5. Interpretation: Deep purple/blue color within 10-30 seconds = Positive. No color change or >30 seconds = Negative.

6. QC: P. aeruginosa ATCC 27853 (Positive), E. coli ATCC 25922 (Negative).

Frequently Asked Questions

Nickel/chrome alloys can cause a false-positive reaction. Use wooden sticks, plastic loops, or platinum.