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Electrophoresis is a laboratory technique performed with the objective of separating molecules according to their size and electrical charge so that diseases can be diagnosed, the expression of proteins is verified or microorganisms can be identified.
Electrophoresis is a simple and low-cost procedure, being used in laboratory routine and in research projects. According to the purpose of electrophoresis, it may be necessary to carry out other tests and exams in order to reach a diagnosis, for example.
What is it for
Electrophoresis can be performed for different purposes, both in research projects and in diagnosis, since it is a simple and low-cost technique. In this way, electrophoresis can be performed for:
- Identify viruses, fungi, bacteria and parasites, this application being more common in research projects;
- Paternity test;
- Check protein expression;
- Identify mutations, being useful in the diagnosis of leukemias, for example;
- Analyze the types of circulating hemoglobin, being useful in the diagnosis of sickle cell anemia;
- Evaluate the amount of proteins present in the blood.
According to the purpose of electrophoresis, it may be necessary to perform other complementary exams for the doctor to conclude the diagnosis.
How it's made
Electrophoresis is a laboratory technique and, to be performed, it is necessary to prepare an agarose or polyacrylamide gel, depending on the purpose of the technique, buffer solution, electrophoresis vat, molecular weight marker and a substance capable of allowing the visualization of samples when exposed to UV or LED light.After preparing and solidifying the gel, the technique can be performed as follows:
- Place each sample in a well of the gel, mixing a small amount of molecular weight marker;
- Place in one of the wells a positive control, which is the substance that is known to be, and in another well, the negative control, to guarantee the validity of the reaction. In both wells, the molecular weight marker must also be present;
- Place the gel in the electrophoresis tank, if not, with the specific buffer solution, and turn on the device so that an electric current is generated capable of generating a potential difference and separation of particles according to their charge and size. The electrophoretic run time varies according to the purpose of the procedure, and can last up to 1 hour;
- View the electrophoresis result using the transilluminator When the gel is placed under UV or LED light, it is possible to visualize the band pattern: the larger the molecule, the smaller it is its migration, getting closer to the well, while the lighter the molecule, the greater the migratory potential.
For the reaction to be validated, the bands of the positive control must be visualized and that nothing is visualized in the negative control, otherwise it is indicative of contamination, and the entire process must be repeated.
Types of electrophoresis
Electrophoresis can be performed for different purposes and, according to its purpose, different types of gel can be used, the most common being polyacrylamide and agarose.
Electrophoresis to identify microorganisms is more common to be performed in research laboratories, however, for diagnostic purposes, electrophoresis can be used to identify hematological diseases and diseases that evolve with an increase in the amount of proteins, being the main types of electrophoresis:
1. Hemoglobin electrophoresis
Hemoglobin electrophoresis is a laboratory technique performed to identify the different types of hemoglobin circulating in the blood, making it possible to identify the presence of diseases related to the synthesis of hemoglobin.The type of hemoglobin is identified by electrophoresis at a specific pH, ideally between 8.0 and 9.0, and a pattern of bands is verified that can be compared to the normal pattern, allowing the identification of the presence of abnormal hemoglobins.
What it is used for: Hemoglobin electrophoresis is used to investigate and diagnose diseases related to hemoglobin synthesis, such as sickle cell anemia and hemoglobin C disease, in addition to being useful in the differentiation of thalassemias. Learn how to interpret hemoglobin electrophoresis.
2. Protein electrophoresis
Protein electrophoresis is a test ordered by the doctor to assess the amount of circulating proteins in the blood and thus identify diseases. This test is performed using a blood sample, which is centrifuged to obtain plasma, which is the part of the blood made up, among other substances, of proteins.
After electrophoresis, a pattern of bands can be visualized and, later, a graph showing the amount of each protein fraction, which is essential for diagnosis.
What it is used for: Protein electrophoresis allows the doctor to investigate the occurrence of multiple myeloma, dehydration, cirrhosis, inflammation, liver disease, pancreatitis, lupus and hypertension according to the band pattern and the graph presented in the exam report.
Understand how it is done and how to understand the result of protein electrophoresis.