Capillary Electrophoresis – Applications and Procedure

What is Capillary Electrophoresis?

It is an analytical method used to separate ions according to their electrophoretic mobility using an applied voltage. There are various factors that can significantly affect electrophoretic mobility such as:

• Charge of the molecule
• Viscosity
• Radius of the atom

The rule of the thumb is that the greater the field strength the faster the mobility. There are different types of electrophoresis but the most predominant one is the capillary electrophoresis because it yields faster results and provides a high-resolution separation. ^{(1, 2, 3, and 4)}

Image 1: The image above shows how capillary electrophoresis takes place.

Picture Source: wikimedia.org

 

What is the basic principle of electrophoresis?

Electrophoresis is a method in which the sample ion moves through the influence of applied voltage. It pertains to the migration of charged ions in the electric field. In a given solution, the electric current flows between the electrodes and carried by ions.

Looking at the principle, the charged molecules are placed in the electric field and migrate towards the pole of either positive or negative charged. A nucleic acid has a consistent negative charge imparted by the phosphate backbone and migrates towards the anode. Hence, the force will accelerate the movement of protein towards the cathode or anode depending on its charge. ^{(2, 3, and 4)}

To sum up the process, the capillary is filled with a conductive fluid with designated pH value. The conductive fluid will serve as the buffer solution in which the sample will be separated. A sample is placed in the capillary through a pressure injection or through electrokinetic injection.

A high voltage is placed over the capillary which will enable the sample to move through the capillary at varying speeds. The positive components travel to the negative electrode while the negative ones travel to the positive electrode.

Image 2: The principle of capillary electrophoresis as shown in the image where positively charged ions are called the anode and the negatively charged ions are called the cathode.

Picture Source: ytimg.com

What is the importance of electrophoresis?

Electrophoresis is a method used by molecular biologists. It pertains to the migration of a charged molecule through the restrictive matrix/gel drawn by an electrical force. Through electrophoresis, laboratory professionals can identify organic molecules and study them for biomedical analysis. The significance of electrophoresis includes the following:image

Antibiotic testing

electrophoresis plays an important role in testing antibiotics. Antibiotics are needed to help combat diseases and infections. What electrophoresis does is it separates the antibodies present in antibiotics from different types of impurities. Through electrophoresis, the researchers can check the concentration of antibiotics leading to an accurate dosage.

DNA analysis

Electrophoresis is known for its usefulness in DNA analysis. With gel as a medium, the researcher can flake DNA into segments with the help of an electrical charge and retain the molecules in their position the moment the charge is removed. Through electrophoresis, the researchers can check high resolution molecules. Hence, it would be easy to analyze the structure of DNA.

Vaccine testing

The development of modern vaccines is made possible with the aid of electrophoresis. The process of electrophoresis is useful in checking the vaccines’ purity and concentration. It enables the testing of different vaccines with different types and levels of antibodies. ^{(5, 6, and 7)}

Protein and antibody analysis

With this type of electrophoresis, a special process called immunoelectrophoresis is used, which enables researchers to check proteins and antibodies interactions. Through immunoelectrophoresis, different types of immune conditions can be detected such as kidney-related diseases and multiple sclerosis.

It also helps detect the interaction of antibodies with unusual proteins that might be present in the samples. Hence, enabling to find the best treatment, cure, and management remedies for autoimmune diseases. ^{(1, 5, and 7)}

What is capillary electrophoresis used for?

Capillary electrophoresis is mainly used in forensic science, specifically in the development of methods for DNA fragment amplification and detection with the use of polymerase chain reaction.

• Capillary electrophoresis is used by a forensic biologist for the typing of STR from the samples (biological) to come up with a profile from highly polymorphic genetic markers, which are distinct in every individual.
• Capillary electrophoresis helps detect the specific mRNA fragments to detect biological fluid of tissue origin in a given forensic sample.
• Capillary electrophoresis is also useful in ink analysis, which is becoming more important as more and more cases of counterfeiting of documents are detected using inkjet printers. Through capillary electrophoresis, fraudulent documents and counterfeit banknotes will be detected.
• A special type of capillary electrophoresis known as affinity capillary electrophoresis is conducted to understand the interactions of protein-ligand. It is commonly used in pharmaceutical companies and life sciences. ^{(7, 8, 9, and 10)}

Types of capillary electrophoresis

1. Capillary isoelectric focusing –  It is used to separate peptides and proteins. The two determining factors are the charge of protein groups and the pH of the solution. As the pH is altered, the charge also changes.
2. Micellar electrokinetic capillary chromatography – It uses a surfactant to form micelles; fat molecules that form a sphere in aqueous solution. Micelles have polar negativity, which pulls towards the positive pole. Molecules that hate water will aggregate within the micelle while water-loving molecules move quickly through the solution.
3. Capillary zone electrophoresis – Of all capillary electrophoresis methods, the capillary zone is the most common. It is also known by the name free solution capillary electrophoresis. In this type of separation method, the ratio of charge particles to mass is taken into consideration. Hence, the larger the ratio the quicker the separation will be.
4. Capillary electrochromatography – It is a combination of capillary electrophoresis and liquid chromatography. It has three parts – open tubular capillary electromyography, the CEC with some columns, and CEC with monolith.
5. Capillary gel electrophoresis – it requires a regular field strength and is dependent on the pH of the buffer solution. This type of capillary electrophoresis is ideal for macromolecules like proteins and DNA. ^{(3, 5, 7, 9, and 10)}

References

1. https://www.news-medical.net/life-sciences/Types-of-Capillary-Electrophoresis.aspx
2. https://chem.libretexts.org/Bookshelves/Analytical_Chemistry/Supplemental_Modules_(Analytical_Chemistry)/Instrumental_Analysis/Capillary_Electrophoresis
3. https://link.springer.com/chapter/10.1007/978-1-59259-120-6_2
4. https://en.wikipedia.org/wiki/Capillary_electrophoresis
5. https://www.intechopen.com/books/electrophoresis/method-development-by-use-of-capillary-electrophoresis-and-applications-in-pharmaceutical-biological
6. https://www.news-medical.net/life-sciences/Types-of-Capillary-Electrophoresis.aspx
7. https://www.sciencedirect.com/topics/food-science/capillary-electrophoresis
8. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4551542/
9. https://www.labcompare.com/10-Featured-Articles/133264-Capillary-Electrophoresis-Systems-for-High-Resolution-Separations/
10. http://www.princetechnologies.eu/products/ce-systems/ce-technologies/ce-introduction/