About this test
What is the purpose of the test?
The purpose is to see if the microbe can use the amino acid arginine as a source of carbon and energy for growth. Use of arginine is accomplished by the enzyme arginine dihydrolase .
How is arginine dihydrolase activity determined?
A medium containing arginine and a pH indicator is used. When arginine is used, the pH of the medium rises and the indicator changes color.
What medium is used?
The medium used is arginine dihydrolase broth . The medium is a nutrient broth to which 0.5% arginine is added. An important component of the medium is a modest amount of glucose, necessary for the process to proceed. The pH indicator brom cresol purple is purple at neutral or alkaline/basic pH but turns yellow at pH <5.2.
How is the test performed?
An inoculum from a pure culture is transferred aseptically to a sterile tube of arginine dihydrolase broth. The inoculated tube is incubated at 35-37 C for 24 hours and the preliminary results are determined. The microbe must first use the glucose present to cause the pH to drop. This is indicated by a change from purple to yellow . Once the medium has been acidified, the enzyme arginine dihydrolase is activated. The culture is incubated an additional 24 hours at 35-37 C to allow the microbe to now use the arginine . The final results are then obtained by observing the tube at 48 hours. Change back to purple from yellow indicates a positive test for arginine dihydrolase . Failure to turn yellow at 24 hours or to revert back to purple at 48 hours indicates a negative result.
What reagents are needed?
Performing this test in the VUMIE Online lab
Inoculation of Medium
1. Select the arginine dihydrolase broth medium.
2. Complete the process of a tube-to-tube aseptic transfer to inoculate the medium.
Incubation of the Inoculated Medium
3. Place the inoculated tube into the 35-37 C incubator.
4. Press the New Day button to move forward 24 hours.
Determination of Test Results
5. Incubate this test for 24 hours. Make note of the color, then incubate for an additional 24 hours.
6. Retrieve the incubated culture from the incubator.
7. Observe test result. If the test protocol was followed as described above, the culture will have changed to yellow due to acids produced in the first 24 hours and then reverted to purple at the 48 hour mark as the amino acid is used. Both color changes must occur for the test to be positive. Any other result is considered negative.