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Wednesday, August 4, 2021

Electrophoresis



ELECTROPHORESIS:

Under the influence of an electric field, electrophoresis separates macromolecules in a gel or fluid based on their binding affinity, size, and charge. In 1807, Ferdinand Fredric Reuss was the first to detect electrophoresis. Procedures like Anaphoresis and Cataphoresis are included in the basic categorization of electrophoresis. The electrophoresis of negatively charged elements known as anions is known as anaphoresis. Cataphoresis, on the other hand, is the separation of cations or positively charged particles via electrophoresis. The method may be used to analyses and separate biomolecules such as plasmids, proteins, nucleic acids, DNA, and RNA.

Electrophoresis Principles of Operation

In an electric field, charged macromolecules migrate in the direction of the positive or negative pole. The charge of the macromolecules determines the mobility in the end. Because nucleic acid is a negatively charged particle, it prefers to travel in the direction of the anode in this scenario. There are two types of electrophoresis procedures. Capillary electrophoresis and slab electrophoresis are the two methods. Proteins that are negatively charged will migrate towards the anode, whereas proteins that are positively charged will move towards the cathode. Scientists can simply measure the travelled distance and apply logarithms to determine the size of the particles since smaller molecules move quicker than bigger ones.

Different Electrophoresis Methods

Supporting material is included with modern electrophoresis equipment. This supporting medium acts as a physical barrier, allowing charged macromolecules to be separated. This physical support performs two critical functions: molecular sieving and macromolecule adsorption for separation. Agar, agarose, starch, and polyacrylamide are some of the most widely used supporting media. The electrophoresis technique is divided into two types based on whether the supporting medium is available or not: capillary electrophoresis and slab electrophoresis.

Electrophoresis in Capillaries (CE)

You may separate charged particles using this separation approach by looking at their different migration speeds in the electric field. It is entirely dependent on the particle's size and charge.

Electrophoresis in a Slab (SE)

It is a widely used technique for separating proteins. The 1D format allows you to evaluate several samples at once. Though CE requires specialized equipment, SE does not. These two types of electrophoresis techniques can be further subdivided into several sorts of electrophoresis techniques. Techniques like paper electrophoresis and gel electrophoresis are included in the capillary electrophoresis category.

Zone electrophoresis, Isoelectrofocusing, and Immunoelectrophoretic are all sub-categories of slab electrophoresis.

Below, we'll go through the specifics of each of these distinct forms of electrophoresis:

1.      Gel Electrophoresis:

In the majority of experimental settings, it is one of the most popular electrophoresis techniques. Gel electrophoresis may be divided into three types. Starch gel electrophoresis, polyacrylamide gel electrophoresis, and agarose gel electrophoresis are the three types of electrophoresis. Potato starch granules are utilized as a supporting medium in the starch gel electrophoresis process. In the case of agarose gel electrophoresis, things are a little different. The support medium is a completely pure polysaccharide with a high molecular mass. Because of its great stability, polyacrylamide gel electrophoresis is one of the most widely used methods. It also works with a wide range of molecular concentrations.

2.      Paper Electrophoresis:

The method is straightforward. The material to be separated was placed on a strip of paper that had been moistened with a buffer solution. This buffer solution is separated into separate tanks, and each end of the paper is dipped in one of them. A separate cathode or anode is also present. After that, an electric current is used. It pushes the sample to travel in the opposite polarity direction of the electrode. The paper is then dried and examined using a sound quality detecting device once the operation is done.

3.      Immunoelectrophoretic:

A separate cathode or anode is also present. The next step is to apply an electric current. It causes the sample to travel in the opposite polarity direction of the electrode. The paper is then dried and inspected using a sound quality detecting device when the operation is done.

4.      Zone Electrophoresis:

Zone electrophoresis, or ZE, is a technique for analyzing nucleic acids, biopolymers, and proteins. This electrophoretic separation technique entails transporting several species in a buffer system while using an electric current. These species will split into well-resolved and diverse peaks due to variations in mobilities.

5.      Isoelectrofocusing

This method works because under an electric field, macromolecules in a pH gradient prefer to migrate towards their pI. The process is performed using IPG strips made of acrylamide gel with large pores to prevent sieving effects. A separate cathode or anode is also present. The next step is to apply an electric current. It causes the sample to travel in the opposite polarity direction of the electrode. The paper is then dried and inspected using a sound quality detecting device when the operation is done.

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