In many ways, biochemical tests play a crucial role in science. Whether it is the identification of bacterial species, Clinical samples of a disease, or research. Without biochemical tests, there will be an enormous amount of data that will be inaccessible. Take for example the identification of gram-negative bacteria from gram-positive bacteria. An effective biochemical test would help narrow down the disease list. Biochemical tests have also played an important role in Molecular biology as methylated DNA is differentiated from non-methylated DNA based on specific tests. That leads to the role of that fragment of DNA in cell activity.
While performing a biochemical test is necessary to understand the different terms related to it. Often, we may come across terms like reagents, medium, nucleophilic and electrophilic reagent. The culture medium with suitable reagents mixture is necessary to perform any test because it copies the in vivo mechanism and provides all the suitable conditions for a cell to grow and evaluate.
Reagent VS Medium
small molecules in nature. The chemical composition of the reagent is known and is optimized for the specific results. When added with the sample they either changes color or produce gases that indicate the presence or absence of the desired molecule. An example is when we want to know the presence of adulterants in milk sample chemicals is added that indicates the presence of a foreign molecule that is normally not the part of the milk.
Similarly, when we talk about medium it carries a whole different meaning. The term is often applied when something is to be grown. Such as when we are testing the antibiotic resistance, the bacteria are grown on a Petri plate that contains a careful mixture of ingredients that are crucial for bacterial growth. Similarly, when mammalian cells are grown, they need a special environment and carefully crafted environment to help them grow outside the body. These cells are grown in special liquid media with many known and unknown ingredients. So, we can say that the difference between a reagent and medium is their composition. While reagents have known composition, the medium has ingredients that are unknown and often not entirely understood for their function.
Chemical VS reagent
A chemical is usually a single substance, for which we can measure and record physical and chemical properties. For example, benzene, or sulfuric acid. Individual chemicals may be used as a reagent. while a reagent takes part in a chemical reaction and is converted to something else. The term reagent can also be used for a common mixture such as a solution of the reacting chemical. For example, NaOH is solid but is often used as a solution in water, and the solution may be called a reagent.
Nucleophilic VS electrophilic
There are some other terms we come across like Nucleophile reagents and electrophilic reagents. As the name suggests the Nucleophile are those reagents that are attracted toward the positive charge. These reagents have extra electrons to donate thus they act as a negative center. Similarly, the electrophiles are those reagents that are more attracted to the negative charge and have a positive charge and are deficient in electrons. These two types of reagents are used to indicate the end-product and identification of the nature of the molecule that we are trying to detect in a sample. An example is a nucleophile reagent that is there during the DNA replication process. Similarly, the reagents used in PCR also act as nucleophiles and electrophiles for different molecules.
The TSI test is another example of the biochemical tests and how they are important in differentiating between different types of Bacteria. TSI (triple sugar Iron) media is a combination of three different sugars and iron. The three sugars are added to differentiate between gram-negative, gram-positive bacteria, and anaerobes. Different bacteria have different abilities to digest sugars. The three sugars are Sucrose, lactose, and glucose. Depending on the color produced in the media we can predict the type of bacteria. Thus, an important tool for the investigation of disease-causing bacteria.
Super optimal broth(SOB) is a nutrient-rich media use for the microbial cultures, Mostly for the transformation of competent cells. It was developed by the American biologist Douglas Hanahan in 1983. Super optimal broth with catabolic repression(SOC) is the SOB with added glucose. Catabolite repression allows micro-organisms to adapt quickly to a preferred (rapidly metabolizable) carbon and energy source first. This nutrient-rich broth consists of peptides, amino acids, water-soluble vitamins, and glucose in a low-salt formulation. It is used mainly in the recovery step of Escherichia coli competent cell transformations. The use of SOC maximizes the transformation efficiency of competent cells. SOC is the adjusted form of commonly used LB media(lysogeny media).
Reagent medium original color test
Many biochemical tests give results based on changes to color in the reaction media. Here are a few examples that include the Iodine test, urease test, methyl red test. These tests are used to differentiate organisms based on their ability to metabolize the regents present in the media. They can also be used as part of the identi?cation of several microbial genera and species.
Urease test: Urease test is a common diagnostic test for the rapid diagnosis of Helicobacter pylori. It is a rapid, cheap, and simple test that detects the presence of urease in or on the gastric mucosa. Hydrolysis of urea produces ammonia and CO2. The formation of ammonia alkalinizes the medium, and the pH shift is detected by the color change of phenol red from light orange to pink. Rapid urease-positive organisms turn the entire medium pink within 24 hours.
Iodine test: Iodine test is also a rapid test used for the presence of starch. When a solution of triiodide anion is added, it changes to deep blue or black color indicating the presence of starch. The intensity of color changes if any other substance present in the tested solution.
Methyl red test: Methyl Red (MR) test determines the ability of a microbe to performs mixed acids fermentation when supplied with glucose. The type and quantity of acid produced differ from species to species and depends on the specific enzymatic pathways present in the bacteria, which helps to determine the various bacterial classification. When methyl red is added to the culture medium, color changes to red indicate the positive test results.
These are the few examples from a long list of tests based on the reagent media color change. They are advantageous over other tests as time saving and cost-effective.