Hepatitis C - infection of the liver caused by the Flaviviridae HCV virus, consisting of one or more RNA molecules. As a rule, a number of diagnostic measures are carried out to establish hepatitis C in a patient. PCR is an analysis that accurately confirms the diagnosis. Often, the conclusion is provided by doctors when the patient already has the first signs of the disease.
What is hepatitis C
The defeat of the liver by a viral infection, transmitted through blood or sexually, is called hepatitis C in medicine. The causative agent is an RNA-containing virus of non-cellular infectious agents of the Flaviviridae family. The infection can be in the body for a long time without manifesting itself. Therefore, only when a microorganism is detected using a blood test by PCR RNA, hepatitis C (as a diagnosis) is considered reasonable.
Flavivirus is not produced in artificially grown isolated cells. In the process of reproduction, the infectious agent creates immunologically different modifications. These factors prevent the body from giving an appropriate protective response, and specialists are having difficulty developing an effective vaccine.
The virus is transmitted in a parenteral manner. For infection it is necessary to get it in a sufficient amount directly into the blood.
What tests do you have for hepatitis C?
Viral liver pathologies are diagnosed using laboratory tests. Clinical studies of hepatitis C are carried out by the method of studying foreign antigens or protein compounds (antibodies) that are foreign and dangerous for the body in the patient’s biomaterial, mainly in the blood.
- Enzyme-linked immunosorbent assay (ELISA). The method is based on the detection of IgM antibodies in the blood. Immunoglobulin is considered the largest and is a pentamer. It manifests itself in the primary protective response of lymphocytes to an unknown foreign substance.
- Radioimmunological analysis (RIA) is a quantitative determination of IgM immunoglobulin using a labeled radioactive isotope of iodine.
- PCR of hepatitis C - determination of viral RNA in the biomaterial (blood). This analysis reliably confirms the diagnosis.
A test for immunoglobulin G is not reliable. Its presence in the blood serum can speak not only about the presence of flavivirus, but also about another transmitted infection that has the same pathogen.
What determines the analysis of PCR
The polymerase chain reaction (PCR) is based on the identification of DNA regions specific for certain pathogens in samples of research material (epithelium, blood). The analysis makes it possible to determine the pathogenic smallest organisms (viruses) by detecting their RNA or DNA in biological material.
The laboratory procedure for the detection of hepatitis C PCR antibodies is carried out in 3 stages:
- Selection. The biomaterial under study is purified from impurities and DNA is obtained in the mobile phase of the chromatographic column.
- Amplification. The analysis is carried out in a device-thermostat amplifier. He with a certain cyclic heats and cools the tubes. For one study, up to 35 cycles are produced. The result is a sufficient number of DNA fragments to identify, identify and evaluate the pathogen.
- Electrophoresis The obtained fragments are placed in a gel with different agarose saturation and electrophoresis is carried out. The resulting electrophoregram is analyzed on a computer.
Not only a standard PCR study is used, but also Real Time PCR analysis. This is a real-time diagnosis. The method allows for a qualitative and quantitative analysis, which makes it possible to identify the formation and dynamics of the disease, as well as prescribe therapy aimed at eliminating the cause of the pathology. Real-time PCR is also used to determine the pure culture of infectious agents (genotyping).
How does an analysis for polymerase chain reaction
For diagnosis, any human biological fluid can be used. The hepatitis C test material is usually blood.
Sampling for diagnosis is carried out in the treatment room. To take blood, use disposable tubes with a substance that inhibits its coagulability. Measures aimed at preventing the spread of pathogens help to avoid the entry of another strain of microorganisms from the outside.
Hepatitis C PCR blood is donated in the morning on an empty stomach. They try to immediately deliver the sample to the laboratory. Storage of biomaterial is allowed at temperatures from +4 to +8 degrees Celsius. The containers are labeled and provided with direction. Analysis results can be obtained after 48 hours, sometimes earlier.
Types of PCR
The range of applications of the polymerase chain reaction is quite wide. Infectionists determine by hepatitis B PCR, diseases transmitted by blood-sucking insects, AIDS, tuberculosis. In oncology, using this method, tumor cells are detected in the early stages.
There are about fourteen types of analysis. The use of one kind or another depends on the scope and end results of the analyzes. Some types of PCR are indispensable where the result is needed within 20 minutes.
For the diagnosis of hepatitis C, 3 types of polymerase chain reaction are used:
- A qualitative assessment can be either positive indicating the presence of infection in the body, or negative - the absence of flavivirus.
- Real-time PCR (quantitative analysis) - determines the quantitative RNA of the pathogen in IU / ml.
- Genotyping - an analysis that identifies a variety (genotype) of a virus.
To accurately identify and determine the disease, followed by the appointment of optimal therapy, all three types of studies are used.
Qualitative analysis of polymerase chain reaction
The analysis is prescribed without fail to all who ELISA revealed antibodies to HCV. Qualitative PCR for hepatitis C is a standard sensitivity test. The method is aimed only at detection, no calculation or isolation of other substances is performed.
To determine the antibodies using special test systems with a sensitivity threshold of at least 50 IU / ml. If antibodies are detected, other refinement tests are prescribed. If the result is negative, no further studies are carried out.
In some cases, a false negative result may be due to incompetence of laboratory staff or low-quality reagents. For insurance, it is better to retake the analysis elsewhere.
Quantitative PCR
The method is used to directly study the number of flavivirus per reaction cycle. DNA fragments used for hybridization or fluorescently-labeled primers are used for accurate measurement. There is an economical detection option using SYBL Green. The dye is wedged into a small groove of the DNA and turns blue when irradiated with a laser.
The concentration is determined by the device-amplifier in digital terms. In different laboratories, the indicators may vary slightly, so they need to be compared with reference indicators.
Quantitative PCR of hepatitis C helps to choose the optimal dosage of drugs and determine the duration of therapy. The frequency of the study depends on the stage of the disease, the type of genotype and the prescribed course of treatment.
Genotype determination
Hepatitis C virus has a variable genetic structure. Numerous modifications make it impossible to create a vaccine, and also complicate therapy. In total, 11 genotypes and 100 subtypes were identified and fixed. In countries of the former USSR, in individuals infected with hepatitis C, PCR mainly reveals genotypes 1b and 3.
In the presence of any of the genotypes, cirrhosis or liver cancer can frolic. Therefore, it is so important to detect the virus on time.
For some patients, certain drugs designed to fight HCV are toxic. Genotyping allows you to determine the type of protein and prescribe an effective drug.
In the results of typing tests, there is a figure with a lowercase Latin letter indicating the genotype of the virus. If HCV is detected, but not typed, this means that a person has a genotype that is unusual for a certain geographical area.
Analysis results
The doctor decides the results. Only the data of several studies (combined with an anamnesis and examination) can give a general real picture of the medical history.
- In a healthy person, a test for a qualitative reaction in a biomaterial does not reveal anything. If the value "detected" is indicated, the infection is confirmed, and the patient needs further diagnosis with subsequent treatment.
- Determining the number of infectious agents makes it possible to assess the viral load on the body. In normal quantitative PCR of hepatitis C, the pathogen is not detected. An indicator up to 8 * 10 ^ 5 is regarded as a low load and with proper therapy guarantees a favorable outcome. Higher values require an in-depth examination and determination of long-term therapy, for a positive outcome of which no one can vouch.
- Positive genotyping results indicate which genotype is recognized. A negative result indicates either the absence of flavivirus or the presence of a genotype that is not characteristic of the region.
What a positive PCR analysis indicates
Any serious disease requires a comprehensive diagnosis. Positive hepatitis C PCR confirms the diagnosis, but predictions can only be made after additional laboratory and instrumental studies.
Detection of the virus does not give a complete picture of the pathology. It is necessary to identify its type and character, to determine how it affects the liver and other organs. Early detection of HCV often has a favorable therapeutic outcome.
Negative PCR with positive ELISA
When observing symptoms characteristic of liver damage, there is a high probability that the infection has already entered the body. Therefore, you should contact a hepatologist or infectious disease specialist. A specialist in the course of interrogations will collect an anamnesis and prescribe the necessary examination.
If, according to the results of hepatitis C studies, PCR is negative, and the enzyme immunoassay is positive, then this indicates the presence of antibodies to flavivirus in the blood. Usually this happens when the infection enters the body in a small amount, so the immune system coped with it on its own. But such people are still considered infected and must undergo an examination every 6 months. If with such test results a person is denied a free medical examination, to maintain health it is better to take PCR for a fee, and if positive, contact a specialist to make a diagnosis and prescribe treatment.
Advantages and disadvantages of the method
When diagnosed with hepatitis C, PCR has several advantages:
- Detection of the pathogen in the early stages.
- Exact definition of a virus.
- The efficiency of the diagnosis.
- Minimum probability of error.
- High sensitivity.
The disadvantages of the method:
- High cost of analysis. The test requires expensive equipment, reagents and highly qualified medical staff. All together adds up to a considerable amount.
- Strict requirements for the transportation of biomaterial.
Treatment of liver inflammation
An analysis of PCR for hepatitis C virus is considered fundamental for diagnosis, but not definitive. For therapy to be effective, a number of additional laboratory and instrumental studies are required. After passing all the diagnostic measures, the doctor prescribes treatment:
- Prescribed diet number 5.
- Alcohol is completely ruled out.
- Joint reception of "Interferon" and "Ribavirin" for 25 days.
- Coursework hepatoprotectors "Essentiale", "Karsil", "Phosphogliv".
- In special cases, discrete plasmapheresis is used.
Medications prescribed by the hepatologist are used regularly without changing the dosage. The duration of medication is determined by the doctor after passing control tests. After completing the course of therapy, you must visit the doctor every six months.
PCR diagnostics can identify the causative agent of hepatitis C in the early stages. This contributes to the effectiveness of therapy and preventing the transition of the disease into such dangerous forms as cirrhosis and hepatocellular carcinoma.