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Antibiotic Assay To Determine The Potency Of Antibiotics

Submitted on March 27, 2012

An antibiotic assay is used to determine the potency of the antibiotics using an assay technique. The potency of the antibiotic can be examined by creating a suitable environment. Its inhibitory effects are then studied on the respective microorganisms.

An antibiotic sensitivity assay tests the efficacy of a drug in inhibiting the growth of microorganisms that causes a disease. Antibiotic assays are carried out on the infecting organism that is found in clinical specimens of body fluids such as blood and urine collected from patients seeking treatment. Isolation and identification of the pathogens responsible for the disease is necessary before selecting the appropriate antibiotic for testing. Once isolated from the patient's specimen, the organism is cultured in a medium with a nutrient broth at pH 7.0, in most cases. Then, the culture is inoculated with a suitable solution of the antibiotic to be used to fight the disease, and its effectiveness in inhibiting the growth of the microorganism is recorded.

Antibiotic Assay In Body Fluids

The Stokes method tests sensitivity to an antibiotic by using an inoculation of a controlled organism on a part of the plate, while the rest of the plate is coated with the test organism. Disks placed at the interface allow the comparison of the zones of inhibition.

Since the antibiotic is known to work on the control organism, its effectiveness against the test organism can be measured easily by comparing the two zones. E test uses a process of dry chemistry that is based on impregnating plastic strips with various concentrations of an antibiotic. Antibiotic assay in body fluids is also done for cancer patients who undergo chemotherapy due to the increased risk of infections to ensure the adequacy of the antibacterial drugs used and to prevent toxicity due to overdose.

Different Methods For Conducting Antibiotic Assays

The type, pH, and volume of the medium used for growing the culture, the age of the culture, the size of the inoculums are all important factors that have a bearing on the results of the antibiotic assay test. Even the temperature during incubation and the period of incubation may affect the outcome of a microbial antibiotic assay.

The techniques adopted to test microbial sensitivity to a particular antibiotic are called antibiotic assay methods. The test-tube dilution method and the use of antibiotic assay discs are two of the most popular methods in use. In the test tube dilution method, to check the sensitivity of a microorganism to an antibiotic, a serial solution of the antibiotic in a suitable medium is prepared. Each test tube with a different concentration of the antibiotic is inoculated with a specific number of microorganisms and incubated for a specified period of time. The test tube with the lowest concentration of antibiotic, which inhibits the growth of the microorganism as shown by the absence of turbidity, is identified as the one with the minimal inhibitory concentration.

The antibiotic disc assay is performed by uniformly coating an agar plate with the organism being tested and a paper disc soaked in a suitable concentration of the antibiotic solution. This is placed on the surface of the plate. Since the growth of the organism and the introduction of the antibiotic start at the same time, it creates is a circular zone of inhibition around the disc where the antibiotic stops the growth of the organism. The diameter of this zone depends on the concentration of the antibiotic in the disc and its effect on the growth of the microorganism. Multiple discs containing different antibiotics or one antibiotic in different concentrations can be used to compare and choose the antimicrobial agent that is the most effective in the least concentration to treat the disease. It is important to identify the right concentration of the antibiotic as with repeated use or insufficient dosage the bacteria are capable of developing resistance to the drug. For routine tests, usage of disc diffusion method is more common.

There are two different methods of conducting antibiotic assays. The first method is conducted by using the cylindrical plate method, and the second is the tube assay which uses a turbidimetric tube.

Since these assays may give out differing results, it is important to standardize the medium of the culture and control testing conditions. If not done perfectly, the results may not be standardized and therefore unsatisfactory. Here's how these tests are performed:

Cylinder Plate Method

This test is used to determine the sensitivity of an organism. This organism is usually isolated from a patient before the tests are begun. This is a method in which the antibiotic solution is diffused from a cylinder which is placed on an inoculated surface. After incubation, the diameter of the inhibition zone is measured. This diameter depends on the concentration of the antibiotic used and its specific activity. This method is popular and is therefore used in the commercial preparation of antibiotics.

The result of such an assay can help a physician determine the kind of drugs that need to be prescribed. If the organism is susceptible to the antibiotic in this test, it is highly likely that it will also be killed off when the antibiotics interact with the microbe in the blood stream.

Turbidimetric Tube Method

In this method, a uniform solution of an antibiotic is made. The microbial culture is added to the fluid. The biggest advantage of this method is that it requires a relatively shorter incubation period. However, there is also a big disadvantage. The presence of foreign material that may be inhibitory to the growth of microbes may influence the results of this assay. This method is therefore appropriate when the samples are clear.

Antibiotic protection assay is another assay that might be done in order to check the ability of the microbe to invade the epithelial cells and cause harm. Such a test can help the doctors determine the course that will be taken by a disease that is caused by microbes. It can also help the doctors determine the kind of treatment that should be given to the patient for the particular microbe. Additional tests may be required with the antibiotic protection test.

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