Although it may seem intricate at a first glance, capillary electrophoresis is an instrumental technique with very simple principles. Minute capillaries are used for the electrokinetic separation of analytes in an electrolyte solution under the influence of a given electric field.
Capillary electrophoresis is a superior form of electrophoresis. It reduces the problem of sample overheating. This is due to its high surface to volume capillary tube ratio, which without much difficulty radiates heat. Moreover, it allows for the detection of migrating molecules, by illuminating the tube and detecting the light from the opposite side. The preceding two factors, which were once limitations with traditional electrophoresis, have been effectively solved with capillary electrophoresis.
The molecules are separated in a system with several main components: a capillary tube, electrodes, high voltage electrical source, detector, device for the interpretation of data and small sealed glass capsules (ampoules) for the sample, source and destination.
The capillary tube together with the source and destination ampoules are filled with an electrolyte solution. The capillary tube is then inserted into the sample vial and the sample may be acquired through capillary action, which essentially is the capability of the solution to flow through the capillary without any influence from external forces, including gravity.
External influences may be used in the process of acquiring sample. These include applying pressure, siphoning and electricity.
Once the sample has been introduced, the capillary is then added to the source vial, where the migration of the molecules may begin with the application of an electric filed between the source and destination vials. On the basis of the electro-osmotic gradient created and their electrophoretic mobility, the samples migrate and separate as they do so.
The migration time and electrical field strength are imperative parameters for determining the electrophoretic mobility. In addition to these, the rate of the electro-osmotic flow of the electrolyte solution or buffer used, which is usually greater than that of the sample, has an impact on the sample’s migration velocity.
The rate at which the particle moves during the migration is directly proportional to the strength of the electrical field, the stronger the electrical field, the faster the particle moves.
The samples are then ultimately detected at the opposite end of the capillary tube that they were introduced into. The detection of the sample can be done with a variety of devices. The most commonly used mode of detection is done with ultraviolet light.
The information acquired on detection is sent to a computer or any data processing device, which interprets the data and produces a diagram, which is a function of time that illustrates the separated chemical compounds in relation to each other.
VIDEO Applications of capillary electrophoresis
Capillary electrophoresis has a wide range of biomedical applications. These applications include identifying unknown compounds in a sample, separation of biomedically important proteins, the diagnosis of metabolic disorders and the monitoring of drugs in body fluids among others.
This powerful method for the separation of compounds has many advantages over other existing separation techniques, such as very little sample amounts and solvents are needed. Moreover, it can be applied in virtually all areas of the biomedical field.