Description
Method of determination Quantification, PCR with real-time detection.
Material under study Blood plasma (EDTA)
Home visit available
Determination of HIV type 1 RNA in blood plasma by PCR with real-time detection. The study is performed on the equipment of the Hoffmann-La Roche company (Switzerland) using a standardized technology with automated sample preparation.
The study of the concentration of HIV type 1 RNA in the blood, which is used to predict and monitor the effectiveness of therapy.
The human immunodeficiency virus (HIV) is the etiological agent of acquired immunodeficiency syndrome (AIDS). HIV infection can be transmitted sexually, through contaminated blood and blood products, or from an infected mother to her fetus. Between 3 and 6 weeks after infection, a non-lasting acute syndrome usually develops, characterized by flu-like symptoms and high levels viremia in peripheral blood. In most cases, this is followed by an HIV-specific immune response and a decrease in plasma viremia, usually within 4 to 6 weeks after the onset of symptoms. After seroconversion (appearance of specific antibodies), clinically stable asymptomatic phase capable of lasting for years. The asymptomatic period is characterized by a low level of persistent viremia in plasma and a gradual decrease in the levels of CD4+ T-lymphocytes, which subsequently leads to the development of severe immunodeficiency, multiple opportunistic infections, oncogenesis, and death. In individuals with established HIV-1 infection, quantitative measurement of the level of HIV-1 RNA in the blood is used to predict and control antiretroviral therapy.
Analytical indicators:
Clinical specificity of the test: 100%, with a confidence interval of 99.6-100%.
Test sensitivity: from 20 copies/ml.
Linear test range: 20 - 1 * 10 7 copies / ml
The subtypes of the M and O group (HIV-1) are analyzed.
Indications for appointment
The test is intended for use in the management of patients infected with HIV-1 in conjunction with clinical findings and other laboratory markers of disease progression. The study is used for prognosis (based on baseline HIV-1 RNA levels) or for monitoring the effectiveness of antiretroviral therapy (based on changes in plasma HIV-1 RNA levels over the course of antiretroviral therapy).
Caution: The test is not intended for screening for HIV-1 blood and blood products or for the confirmatory diagnosis of HIV-1 infection.
Interpretation of results
The interpretation of test results contains information for the attending physician and is not a diagnosis. The information in this section should not be used for self-diagnosis or self-treatment. An accurate diagnosis is made by the doctor, using both the results of this examination and the necessary information from other sources: history, results of other examinations, etc.
Units of measurement: amount of human immunodeficiency virus type 1 RNA detected, expressed in C/ml (copies/ml). One copy of HIV-1 RNA corresponds to 1.67 International units(IU) according to the first WHO international standard for HIV-1 RNA in nucleic acid detection tests (NIBSC 97/656).
Result interpretation:
- "not detected": HIV-1 RNA was not detected or meaning below the limit of sensitivity of the method (20 copies/mL). The result is interpreted as "HIV-1 RNA not detected";
- < 20 C/ml (копий/мл): HIV-1 РНК выявлена в концентрации на пределе чувствительности метода, количественная характеристика в копиях/мл с удовлетворительной точностью невозможна;
- from 20 to 10000000 C/ml (copies/ml): The value obtained is within the linear range, the result is reliable;
- >10,000,000 C/ml (copies/ml): Result is interpreted as: “HIV-1 RNA detected at a specified concentration outside the upper limit of the linear range; the test was performed in a 1:X dilution.
Please note that the timing of PCR studies may be extended when confirmatory tests are carried out.
The human immunodeficiency virus is one of the most terrible diseases of the past and present century. Its study, the search for vaccines and drugs bring certain results. However, to date, specialists in the field of medicine have not been able to completely defeat a dangerous disease. Modern methods diagnostics allow not only to detect the immunodeficiency virus at an early stage, but also to control the course of the disease in the future. In order to decide on the need to apply this or that therapy, an HIV RNA analysis is used. It is also used to detect this disease. It is important to note here that HIV RNA analysis is not universal. It is used only when necessary. How is such a study carried out, in what cases is it used and what is its reliability?
HIV virus by RNA: the essence of the study
Before talking about how HIV RNA is determined, you should find out what this abbreviation means and what role it plays in the body. Ribonucleic acid is a molecule found in all living organisms and some viral infections. The synthesis of this molecule is based on a matrix of deoxyribonucleic acid. Certain enzymes are responsible for these processes. These are ribonucleic acid polymerases.
The essence of this type of research is to check the genetic material of the virus. In order to perform an RNA analysis for HIV, medical professionals use the polymerase chain reaction. PCR allows you to repeatedly increase the volume of biological material used for research, due to which it is checked in several stages using a comparative method. It should be noted that PCR for RNA is used not only for HIV. This technique allows you to identify a large number of hereditary diseases, some viruses and infections. It is also used to determine hereditary genetic pathologies. Sperm, saliva and blood are used as biological material. In the case of the immunodeficiency virus, serum and blood plasma are considered to be the preferred material.
The HIV RNA test is performed in several steps. After the patient's blood is taken, it is sent to the laboratory. Biological material is placed in a special reactor, where it is split. For synthesis, special enzymes are used. Further, under artificially created conditions, two are synthesized from one RNA molecule, four from two, and so on. Upon reaching them the right amount, there is a comparison with different types of viruses. By the way, at the exit from one molecule of ribonucleic acid, several hundreds or thousands of molecules are obtained. HIV RNA ultrasensitive is of several types. It all depends on the purpose of the study.
HIV RNA 20 IU/ml: accuracy and reliability of the method, types
In modern medicine, two methods for the study of ribonucleic acid are used. We are talking about quantitative and qualitative HIV RNA. Both of these methods of research are similar, but they have different goals. HIV RNA quantitative is the definition of viral load. The number of viral molecules in genetic tissue can speak volumes. In particular, at what stage is the disease in this moment. Everyone knows that the immunodeficiency virus has several of them. After the onset of seroconversion, the number of virus cells increases, and its detection becomes possible. On the early stages HIV RNA quantitative, the norm of which is established in the laboratory, allows not only to detect the immunodeficiency virus, but also to decide on the need to take antiretroviral therapy. Having determined the quantitative viral load, the specialist can decide which therapy regimen should be used by the patient.
Neonatal quality HIV RNA is used frequently. It can also be carried out in adults, for example, in order to determine the type of virus. But the cost of such a study does not allow it to be implemented in all medical institutions. Therefore, it is used in pediatrics and neonatology.
The reliability of HIV RNA performed with a sensitivity of 20 IU/ml is high. It is about ninety-one percent. However, in modern medicine there are methods whose reliability is higher. That is why enzyme immunoassay or immune blotting is used to determine the presence or absence of this diagnosis. Their reliability is somewhat higher. It is ninety-eight - ninety-nine percent.
If we talk about the most optimal timing for the test, then doctors prescribe it at least a month after infection. If the main goal is to determine whether the patient has this diagnosis, it is not worth analyzing HIV RNA after 4 weeks, its reliability may be inaccurate. It is better to do this in five to six weeks.
If HIV RNA is not detected, what does this mean? This question is often asked by medical professionals. This may mean that the immunodeficiency virus is not present, or indicates that the test was carried out too early.
RNA detection of HIV in newborns: how effective is it?
The preferred method of testing newborns for the presence or absence of the immunodeficiency virus, if they were born to an infected mother, is the analysis of ribonucleic acid. In this case, HIV RNA 20 IU/ml is produced in the first 10 days of the baby's life. It is also possible to detect the presence of the disease with the help of enzyme immunoassay. However, physicians face another challenge. We are talking about the need to determine which antibodies the baby's body produces. The fact is that a baby born from an infected mother will have them anyway. However, sometimes it's just a defense. immune system and not the fact of infection. The reliability of PCR HIV RNA is checked again after 6 months.
According to WHO statistics, for 40 recent years 25 million people have died from AIDS. The damage from this infection is enormous. Most of the infected turned out to be in Africa, where this infection came from. With HIV, the diagnosis of the disease becomes especially relevant. True, there are no drugs for the treatment of HIV yet, but early therapy can prolong the patient's life and improve its quality.
Diagnosis of HIV infection
HIV infection is determined at different stages by the following methods:
- ELISA - enzyme immunoassay.
- Western blot.
- PCR - polymerase chain reaction.
- Express tests.
The PCR method was developed by the American biochemist Kary Mullis, who received the Nobel Prize in 1983 for this. Today, this method in medicine in the diagnosis of all infections is considered the leading one due to its accuracy and information content. HIV is no exception in this regard.
The essence of the analysis
Every living cell contains RNA and DNA. These nucleic acids are capable of self-copying and self-replication. For each infection, DNA fragments are unique. These fragments of nucleic acids circulate in biological fluids. They are captured and recognized by special equipment - the reactor. This is the basis of the method. The laboratory assistant counts these fragments. The retrovirus HIV is monitored for RNA. Even with single copies of viral particles, PCR can detect and count them.
Venous blood is most commonly used as the test fluid. Special components of the method come into contact with the virus particles and make them detectable, because there is a multiple increase in the found fragments.
In case of HIV result the presence of the virus can be obtained long before the clinic appears. Therefore, due to its high sensitivity, PCR has such a high diagnostic value. A huge plus of PCR is the versatility of the method. The incubation period of infection for PCR is not an obstacle.
Research cost
The PCR method is quite expensive. This is one of its big disadvantages. It requires the latest equipment and highly qualified laboratory assistant. In view of the foregoing, PCR diagnostics are not performed in small settlements. You can take the analysis only in specialized large clinics.
The cost of determining HIV DNA in Moscow clinics is from 2,800 rubles, determining the viral load (viral RNA in plasma) using PCR - from 8,800 rubles, and HIV resistance to protease inhibitors - from 16,500 rubles. As you can see, the prices are quite high. PCR can be done free of charge in public clinics under the MHI policy. It is important that the procedure can be done anonymously. In the registry, the patient receives a number by which he can find out the result. Modern medical centers have personal accounts customers where this data will be entered.
Research objectives
PCR diagnostics for HIV is prescribed in such cases:
- Detection of infection in a child born to a sick mother or carrier to determine intrauterine infection.
- If the ELISA gave questionable results (PCR for HIV helps in this case to make a final diagnosis).
- To determine the quantitative content of the virus in the body.
- With positive immunoblotting, they complement each other with PCR.
- Donor testing.
- For early diagnosis of HIV.
- To determine the effectiveness and resistance to ART.
The advantage of this analysis is that PCR for HIV can be performed even for children under one year old.
The study is often performed not for primary diagnosis, but already in the process of treatment. Primary diagnosis is serological studies (they determine the level of antibodies to HIV). If a false-positive result is repeated - this may be with a low viral load.
PCR diagnoses HIV when there are no antibodies to it yet. The ELISA method in this case will not give an answer.
And PTSR on a HIV will be already positive. But the symptoms of this period of HIV are nonspecific. The patient is first treated for a long time and unsuccessfully for ARVI by a general practitioner. It should be borne in mind that the diagnosis of HIV only on the basis of PCR is not made, it is necessary to pass other comprehensive tests. The PCR method is more often auxiliary for complex cases.
Preparation for analysis
Before taking PCR for HIV, 2 days before the analysis, you should not eat fatty foods, alcohol. It is also better not to strain mentally and physically. If the patient is prescribed a course of immunostimulation, it is stopped 2 weeks before the analysis.
It is best to donate blood in the morning. Other biological fluids of the body (semen, vaginal secretion) can also be examined, but best material is blood. Saliva, sweat, urine and tears are not used, since the content of the virus in them is minimal.
Advantages of PCR for HIV
The advantages of the method are the extremely low probability of a false positive result, the universality of the reaction for any biological fluids of the body. The analysis has a wide range:
- One blood draw can be used to detect different infections.
- The technique is urgent, the result is ready the very next day.
- Reliability ranges from 85 to 98%.
- The presence of HIV can be determined 10-14 days after infection (there are no antibodies at this time yet).
- There are no age restrictions, it can be carried out immediately from the moment of birth.
Cons of the method
The disadvantages of PCR are as follows:
- Expensive analysis.
- Sophisticated medical equipment is needed.
- High qualification of the laboratory assistant and the doctor taking the analysis is required.
- The reaction is highly sensitive, so the error can be 20%.
- A false positive result may be obtained if the patient has autoimmune processes, oncology, and chronic infections.
- Special cleanliness of the laboratory premises is necessary, because the virus can get into the analysis from the air. Then the result will be wrong.
For laboratories that conduct PCR, to improve the quality of diagnostics, special strict measures have been developed for the SanPiN system for internal control. In addition, all the rules for working with this technique must be observed:
- It is required to strictly follow the information on the test tubes.
- Before taking blood, look again and make sure that the analysis is ordered.
- The nurse must make the correct labeling of the tubes.
- The laboratory doctor must correctly carry out all manipulations with the biomaterial in order to prevent cross-contamination.
- The test system must be of excellent quality.
Only if all these conditions are met, the error in the answer can be only about 2% of the episodes.
Analysis duration
Many are interested in how long PCR for HIV will be ready. Diagnosis lasts no more than 8 hours. The patient may receive an answer the very next day. Express testing is performed within 2 hours.
Reliability of PCR
Despite the many advantages, PCR is not considered an ideal diagnostic method. They turn to him if it is necessary to obtain screening tests for the presence of HIV in the body.
When is blood taken for analysis?
How long to take PCR for HIV? A reliable result can be obtained already when taking blood 4-4 days after the alleged infection. After 2 weeks, the reliability for HIV will be 98%, and with a period of 5 days - 80%. The presence of PCR on the result will be reliable, but for an absolutely accurate result, ELISA is also performed.
ELISA analysis will be effective only in the presence of antibodies to the virus, this time can take from 1-3 months to six months. Since ELISA gives a higher probability (98% -99.9%), PCR cannot be called a 100% confirmatory test for the presence of HIV infection. But on the other hand, this is the only technique in which it is not necessary to wait for the appearance of antibodies.
With HIV, PCR can provide information about the effectiveness of ART, the stage of HIV disease and the number of VNs (a quantitative assessment of the presence of HIV in the body). This will indicate the severity and extent of the changes.
It is also necessary to take PCR tests for HIV if there are antibodies in the blood, but their presence does not indicate the reliability of HIV infection clinically.
Pass the analysis not only in case of infection and casual sexual intercourse. Other reasons:
- Planning for pregnancy.
- upcoming operation.
- Casual sex.
- Some professions require this analysis for admission to work (teachers, doctors and other medical personnel).
- Prisoners.
- TB patients.
- Employees of the Ministry of Emergency Situations and the police.
- Returned after vacation from exotic countries (if you want to make sure that there is no infection).
- Prostitutes.
- Foreign students.
- Drug addicts.
Also, certain symptoms in a patient can force an HIV test:
- Sudden weight loss.
- Diarrhea lasting more than 3 weeks.
- An unreasonable increase in temperature for a long time.
- Enlarged lymph nodes.
- Unexplained cause of pneumonia, candidiasis, etc.
Deciphering the analysis is the prerogative of the attending physician, not the laboratory assistant.
PCR or ELISA, which is better?
With RNA, the virus is detected both qualitatively and quantitatively. This analysis clearly identifies a specific pathogen even if there are several cross-reacting pathogens. Biological material can be used even in dried form. The downside is the high sensitivity of PCR, when a false positive result can be given by the presence of even a tiny amount of foreign DNA on the instruments or test tube.
The task and possibility of ELISA is to determine the presence of antibodies to a retrovirus. Although its accuracy is 99%, it is not applicable in the early stages.
Qualitative HIV testing
Conducting high-quality PCR for HIV determines the presence of the virus in the body. The results in this case will look like this: positive, false positive, negative. But this study will not give information about the amount of retrovirus. Such a qualitative analysis is inappropriate when HIV infection has already been detected in the body by other methods.
It is impossible to control the effectiveness of treatment with high-quality PCR.
Quantitative HIV testing
It is carried out only by HIV-infected people in order to count the number of copies of virus RNA in a biological product.
The purpose of such a study is to monitor the ongoing treatment and identify the resistance of the virus to it. Antiviral therapy in such calculations is not blindly prescribed by a doctor, so it will be more effective. Quantitative PCR for HIV infection is done much more often. This analysis is displayed as a copy/ml of blood.
What results can be given:
- There is no or very little RNA of the virus (about 20 copies/ml). There is no definite diagnosis.
- From 20 - to 10 to the 6th degree copies / ml - the diagnosis is reliable.
- More than 10 to the 6th copies / ml - large VN.
Laboratories can perform real-time PCR testing for HIV. This implies the observation and numerical evaluation of the accumulation of PCR products with automatic recording of the results.
PCR for HIV is one of the most reliable methods for making this diagnosis, which is dangerous for human health and life. A doctor may recommend testing for any person who has reason to suspect infection with the human immunodeficiency virus. Often patients are interested in what is the essence of the technique, and what are its advantages and disadvantages.
When and at what time is it worth visiting the hospital to hand over the biomaterial for research?
PCR is a diagnostic approach that has appeared in clinical medicine thanks to molecular biology. It was from this branch of science that research came to medicine. And it began to be actively used for making various diagnoses, including infection with HIV infection.
Everyone knows that any living organism consists of DNA and RNA. The polymerase chain reaction is based on the ability of these nucleic acids to reproduce themselves.
It is important to remember that PCR is able to detect even small concentrations of viral particles in the blood. This happens due to the fact that “fragments” of nucleic acids circulate in the biological fluid, which are captured and recognized by special equipment. This allows you to make a correct diagnosis even if the infection has occurred quite recently.
And the viral particles have not yet had time to multiply in the patient's body. The biological material received from the patient is placed in a special reactor. Most often, venous blood is used.
Special components, interacting with virus particles, multiply the number of detected fragments. As a result, the fragments become detectable, they can be evaluated and conclusions about the infection can be drawn.
Donate blood from a vein according to the standard method.
Do not eat before visiting the doctor, it is better to come in the morning. If the patient is undergoing a course of immunostimulation, it is necessary to interrupt it two weeks before the analysis.
Pros and cons of the technique
Like any other method for diagnosing a particular disease, PCR research has a number of advantages and disadvantages. The advantages of the method include:
- low probability of encountering a false negative or false positive result;
- the ability to use for research not only blood, but also other biological fluids, such as vaginal discharge, sperm, etc. (although, as doctors note, accuracy may decrease);
- the ability to use once taken material for the diagnosis of several diseases at once;
- the speed of obtaining results (if the analysis is urgent, then the results are received the very next day);
- the ability to quickly diagnose an infection and, accordingly, begin treatment of the disease, thus improving the prognosis;
- no age restrictions (blood sampling from a vein can be performed even for a newborn baby).
Naturally, PCR also has disadvantages. These include:
- a rather high price for the study, especially in comparison with the basic screening methods used in most hospitals;
- the need for sophisticated equipment, which is also expensive;
- the likelihood of still encountering false results if the analysis technology is violated, the blood sampling process, or the person suffers from an autoimmune, tumor or chronic infectious disease.
Naturally, it is worth performing a PCR study on the recommendation of a doctor.
The doctor will be able to assess in each case how reliable the results will be. If the doctor deems it necessary, he will advise his patient on another diagnostic method instead of PCR. Or, after receiving the results, he will supplement the plan of diagnostic measures with an additional item.
Many patients are concerned about the question of whether all people undergo a PCR test for HIV. In fact, the diagnosis is not mandatory for everyone. However, there are groups of patients who occasionally have to donate blood for analysis.
- Preliminary diagnostics
- Positive immunoglobulin test
Immunoblotting that gave a positive result for the human immunodeficiency virus must be confirmed using a polymerase chain reaction. Do the same if PCR was performed first. Complementing studies with each other almost completely eliminates the possibility of errors.
- Therapy control
Patients with confirmed HIV status donate blood for PCR quite regularly. This is necessary to monitor the course of treatment and, if necessary, adjust therapy.
- Blood safety assessment
Any donated blood must be tested using polymerase chain reaction.
This excludes the transmission of HIV infection through blood transfusion.
- Assessment of the status of the newborn
If a child is born from an HIV-positive woman, he must also undergo PCR. This is necessary to determine the status of the child and develop tactics for its further management.
Reliability of the study
Many patients are concerned with the question of how reliable the polymerase chain reaction is in the event that it is planned to diagnose the human immunodeficiency virus. Despite the fact that PCR is one of the most modern methods, even it cannot be trusted 100%.
Today, PCR is still not used as a screening method precisely because it can give false positive results.
To avoid this, doctors recommend preparing properly for biomaterial sampling, but even preparation does not always help. For example, a test may give a positive result if a person has a malignant tumor in the body. If he suffers from any chronic infection or, for example, an autoimmune process. Even though there is a possibility of obtaining a false positive result, PCR is used in practice as one of the most reliable methods.
To avoid errors, doctors often recommend that their patients be tested again if the result is positive. Also, in some cases, it is recommended to additionally conduct an ELISA reaction and other studies. Performing a set of tests always gives a more reliable result than performing only one study.
What can the test be used for?
Not all patients know what the polymerase chain reaction is specifically used for in the clinical practice of a doctor.
- It is necessary to detect the disease in the latent period
The immunodeficiency virus can exist latently in the body for a long time.
PCR is the only study that can indicate the presence of a pathogen in the body. Even if there are still very few viral particles.
- Establishment of the genotype
Today, several genotypes of HIV infection have been identified. It is often important to determine which specific genotype infected a person, since the selection of antiviral drugs depends on this.
- Definition of the virus, not antibodies to it
One of the advantages of the analysis is that using the technique, it is possible to detect not antibodies to viral particles, but the virus itself.
Why this is an advantage, many patients are interested. The fact is that if the emphasis is on the detection of the pathogen itself, the likelihood of encountering false positive reactions due to the similarity of various antibodies decreases. As the doctors note, in many respects it was the ability to determine the virus itself, and not antibodies, that made the technique so effective.
What are the deadlines for submitting
Often patients are interested in the question of how long it takes to visit a doctor to donate biological material.
It all depends on what exactly is planned to be determined using the methodology.
It is recommended to search for human immunodeficiency virus DNA if infection is suspected and early diagnosis is necessary. In the blood, DNA on average begins to be determined ten days after the infection has occurred. However, the analysis is recommended to be confirmed by ELISA, first after 4 weeks from infection, and then after 12 weeks.
If the ELISA confirms the diagnosis, the patient's status is regarded as positive.
Research to determine RNA is always quantitative. It is recommended that it be performed if the patient's positive status is confirmed and treatment with antiviral therapy has been started. If there is a lot of RNA in the body, then the acute phase of the disease is diagnosed. If it is not enough, it is considered that the suppression of the pathogen was successful, and the therapy can be changed to a more gentle one.
- Resistance assessment
Some strains of HIV show resistance to the drugs used to treat them. A study is also recommended to assess the likelihood of non-response to treatment. It is performed if there is no improvement after the appointment of therapy. It is also recommended in the acute period of the infectious process.
The human immunodeficiency virus is spreading more and more actively in the human population every year. Anyone can encounter pathology, and therefore it is sometimes vital to know how to correctly diagnose.
PCR is a good diagnostic method if used correctly by a physician. And the main thing for the patient is to follow the doctor's recommendations for preparing for the study, in which there is nothing complicated.
