Microbial Assay

Microbial assays provide information on how antibiotics inhibit microbial growth. The microbial assay is a comparative method. Other compounds whose activity can be studied with a microbial assay include amino acids and vitamins. Measured concentrations of the target compound are compared with the standard concentration of a particular compound with known activity, providing information on the compound under investigation.

Microbial assays are used for many purposes. These include antibiotic sterility testing, testing the mutagenicity or carcinogenicity of chemicals, disease diagnosis, and monitoring using immobilized enzymes. They are also used to evaluate the pharmacokinetic properties of drugs for human and animal use.

What is Microbial Assay

There are several methods of performing a microbial assay. These include the disc diffusion method (otherwise known as the cylindrical cup plate method), the tube assay (or turbidimetric) method, the urease assay, and the luciferase assay. These methods have different uses.

• The cylindrical cup plate method involves using circular plates of nutrient agar inoculated with a susceptible test organism. Some characteristics of the cup plate method include the selection of inoculant concentration to obtain suitable dose-response and sharp inhibition zones at different concentrations of the standard, keeping plates on a flat surface to ensure equal distribution of inoculant, and incubating plates at a suitable temperature over a set time.
• Turbidimetric methods differ from the commonly used cylindrical cup plate assay by using tubes. Controls that are not inoculated with the target compound (usually antibiotics) are used to set the optical apparatus used to measure growth. When the tubes have been incubated, growth is arrested by adding formaldehyde R, or the opacity of tubes can be measured at a set incubation time. Appropriate statistical methods are used to calculate potency.

Microbiological Assay of Antibiotics

The microbiological assay is based upon a comparison of the inhibition of growth of micro-organisms by measured concentration of the antibiotics to be examined with that produced by known concentrations of a standard preparation of the antibiotic having a known activity.

Microbiological Assay of Vitamins

Vitamins and amino acids are essential for the growth of microorganisms. The basis of this assay is to measure the ability of test organism to utilize the substance being assayed under a proper nutritional condition. The organisms require these growth factors (vitamins & amino acids) in micro or nano grams.
The response (growth of test organism) is proportional to the dose (amount of factor) added to medium.

Materials required for Microbial assay of vitamins & amino acids: stock solution. inoculum media. Assay medium. standard curve.
MICROBIALASSAYOF VITAMIN B About VitB12: 12 : Also known as cyanocobalamin/Methyl coblamin. Its a water soluble vitamin. Structure is similar to that of heme where the iron is replaced with cobalt as a centre of molecule. Its main sources are liver, eggs , milk, meat & fish. VitB12 deficiency causes Macrolytic anemia, pernicious anemia. National Research Council, USA recommends a daily intake of about 5mg of vitB12

The test organism selected must be capable of utilizing free cyanocobalamin. E.Coli mutant/Lactobacillus Liechmannii is found to satisfy the requirements. Gram negative bacilli, non-pathogenic, easy to culture & easily available. Isolated from milk, cheese, & other dairy products. Assay is performed by using cuplate method / turbidimetric method

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