Types of serological studies. Serology. Indications for implementation

Object of serology - antigens and antibodies

Serological testing (SI) is based on the ability of antigens and antibodies to bind to each other. SI allows you to accurately identify the presence of a certain virus or bacteria in a patient, blood type and Rh, the body’s reactivity to antibiotics, autoimmune diseases, hormonal disorders, individual protein specificity, and determine the effectiveness of therapy.

An antigen and an antibody fit together like a keyhole and a key; when they meet, they certainly bond. This serological reaction allows the detection of various types of antibodies or antigens.

The ability of the immune system to produce special substances that neutralize harmful particles has been known to scientists for a long time. But it was possible to see the antigen-antibody reaction in laboratory conditions only at the end of the 19th century. It was found that as a result of the formation of the binder, the complexes precipitate, forming flakes or cloudiness. Such reactions are called agglutination and precipitation. In the 20th century, serology, the science of the properties of serum, developed rapidly, and new diagnostic methods were discovered based on antigen-antibody reactions. Today, there are many ways to detect immune complexes, and SIs are an indispensable link in diagnosis, treatment, and the creation of new medications and vaccines.

Indications for analysis.

Serological tests are widely used in obstetrics

Serological studies accompany modern man throughout his life. The first tests to detect certain antibodies are carried out when the baby is still in the womb. A pregnant woman donates blood for various tests to determine a possible Rh conflict or infections that can harm the gestating fetus. When a child is born, he undergoes serological tests to diagnose congenital diseases. Subsequently, tests are taken for the following reasons:

It is advisable to stop taking medications the day before the test

Since serological tests cover a wide range of tests, certain preparation may be required in each specific case, but there are general rules for taking a serological test:

The day before the test, you should limit fatty and fried foods in your diet, and exclude alcohol.
The last meal should be completed 10 - 12 hours before the analysis.
On the day of donating blood, you should not drink coffee, tea or other drinks. You can drink clean water in small quantities.
Drugs are excluded before analysis. If it is impossible to cancel the appointment, you must provide the doctor with a complete list of medications, including vitamins and oral contraceptives.
All other studies or medical procedures are carried out after SI.
It is recommended to stop smoking 12 hours before taking a blood sample.
On the previous day, you should avoid physical and emotional stress, and before the test it is advisable to sit in complete peace for 15 minutes.
SIs are prescribed in the morning and taken on an empty stomach.

Methods of conducting serological research.

A large number of methods can be used in serology

All methods for determining antigen-antibody complexes (Ag-Ab) are based on the use of a known Ag or Ab in the search for an unknown one. If it is necessary to detect Ag, diagnostic immune sera containing specific Ab are needed. If the purpose of SI is to identify Abs, diagnosticums are used - suspensions containing certain Ags.

Serological tests can reflect a qualitative or quantitative result. In a qualitative study, the result can be negative or positive, that is, the desired element is either found or not. The quantitative result is expressed as a numerical value or with “+” signs (usually from one to four).

Indirect hemagglutination is one of the diagnostic methods

Serological research methods:

Agglutination. The binding of Ab to Ag followed by the formation of flakes or sediment. Direct, indirect, detailed, indicative reactions are used to detect antibodies in the serum.
Hemagglutination. Red blood cells with Ag or Ab adsorbed on them are used, which stick together with the corresponding Ab and Ag. The formed complexes fall out in the form of scalloped sediment.
Precipitation. Formation of an Ag-At complex followed by precipitation in the form of a cloud called precipitate.
Coagglutination. Detection of Ag using Ab adsorbed on protein of staphylococcus cells.
Inhibition of hemagglutination. Immune serum antibodies suppress viral antibodies, as a result, viruses lose their ability to stick to red blood cells.
Indirect Coombs reaction. Abs to surface Ags of erythrocytes are determined.
Ringprecipitation. It is carried out on the basis of immune serum with a layer of soluble Ag.
Double radial immunodiffusion. The reaction is based on the diffusion of antiserum and Ag components in agar or agarose wells.
Complement fixation reaction. The connection between Ag and Ab is accompanied by the adsorption of complement; sheep erythrocytes and Abs to them in the hemolytic serum are used as an indicator.
Neutralization. Based on the neutralization of the viral agent by antibodies.
Reactions based on the labeling of Abs and Ags with a fluorescent or enzymatic composition - immunofluorescence (RIF) and enzyme-linked immunosorbent assay (ELISA) methods.

Immunofluorescence method.

"Glow" in a dark-field microscope

This method is based on the use of Abs labeled with a fluorescent substance. Most often, fluorescein isothiocyanate, which has a green glow in ultraviolet rays, is used as a label. The result of the analysis is assessed by the degree of brightness of the glow. A manual method of determination using a fluorescence microscope, an optical microscope with a fluorescent attachment, as well as an automatic method using a microchip cytometer, flow cytometer, or robotic fluorescence microscope can be used.

RIF is carried out by direct and indirect methods. The direct method means the direct binding of sorbed Ag to labeled Ab. The indirect method involves using unlabeled Ab to bind to Ag, and then adding labeled Ab, which avoids nonspecific reactions.

Linked immunosorbent assay.

Enzyme immunoassay as a method of serological diagnosis

ELISA is based on the detection of Ag-At complexes by labeling one of the components. Various enzymes are used as a label, which, as a result of an enzymatic reaction, form a color. There are a variety of ELISA methods, they can be classified as homogeneous and heterogeneous methods.

Homogeneous means that all steps of the reaction occur in solution.

Heterogeneous methods involve phase separation using a solid support.

Homogeneous-heterogeneous methods are based on the fact that Ag-At complexes are formed in solution, and then the solid phase is used for separation.

The principle of the sandwich method

There are many variations of ELISA methods, some of which are presented below.

Sandwich method. A solution with Ag is added to the immobilized Ab, after the formation of complexes, the carrier is washed from excess components and labeled Ab is added. As a result, the Ags are clamped with immobilized and labeled Abs, hence the name of the method.
Non-competitive indirect ELISA. The test serum with Ab is added to the immobilized Ag. When complexes are formed, excess Abs are washed off and labeled Abs are added that can bind to immune complexes attached to the surfaces of the wells.
Competitive direct ELISA. The test serum and a conjugate containing labeled Abs are added to the immobilized Ags. Several types of Ag-Ab complexes are formed: with labeled Abs and with unlabeled Abs, which compete with each other for the connection with Ag.

What infections can a serological test help identify?

Serological diagnostics allows identifying the pathogen

SIs make it possible to identify a certain disease if there is a suspicion of it, that is, the analysis is based on knowledge of exactly what element needs to be looked for.

If the patient has certain symptoms or other studies indicate a disease, a serological test is prescribed.

Serological tests are used to diagnose the following infections:

hepatitis virus A, B, C, D, E;
syphilis;
herpes;
cytomegalovirus;
rubella;
measles;
mumps;
Epstein-Barr;
Helicobacter;
chlamydia;
mycoplasma;
toxoplasma;
Giardia;
helminths;
ureaplasma;
parvovirus;
chickenpox virus;
tick-borne encephalitis virus;
Dengue virus;
whooping cough;
borrelia;
legionella.

Serological tests can detect any infections in the laboratory if the antigen of the infectious agent is available.

Advantages of the serological research method in the diagnosis of infections.

Serological diagnostics - fast and affordable

High sensitivity and specificity.
Wide range of detectable infections.
Early diagnosis of infection.
Possibility of monitoring the development of the disease.
Speed and ease of execution.
Possibility of using a minimum volume of test material.
Research safety.

Nowadays, there are quite a lot of tests that we have to take in life. In this article we will consider such a type of analysis as a serology blood test. This analysis is carried out in order to answer the question of whether a person has any infectious diseases and whether everything is in order with his health. Infectious diseases can also be transmitted sexually, and it is precisely such diseases that can be determined by a serological blood test.

What is a serological blood test?

A serological blood test is a test that can show the presence of infectious, viral and microbial diseases.

Basically, this analysis is based on identifying foreign bodies, namely bad bacteria, infections and other organisms that can cause diseases in humans.

Probably, many people know that when a foreign body, bacteria, or other infection enters the human body, the body actively begins to secrete specific antibodies, which subsequently release protein. It is this protein that helps fight foreign microorganisms. But sometimes our body needs help, because it is a protein that fights to heal and cleanse the body, and a serological blood test allows doctors to determine what disease needs to be treated and how to choose an effective treatment method.

How is a serological blood test performed?

A serological blood test is carried out using two methods.
The first method involves the process of introducing serum into the human blood itself, so that an antibody reaction subsequently occurs, and find out how many pathological cells there are in the body.

This method is provided to determine the disease itself, what it threatens and to choose the appropriate treatment tactics.

The second method prefers the process of introducing specific antibodies from human blood into serum. This method is used only in individual cases, when the disease has already been determined, but medical specialists simply need to find out the number of antibodies.

What features should be taken into account when taking a serological blood test?

The fact is that such products can significantly affect the composition of the blood; everything we eat can deposit sediment in our blood, and it is because of this that we need to adhere to a 3-day diet in order for the body to cleanse itself a little before the analysis.

Under no circumstances should you take a blood test for serology when it contains alcohol, as you can not only spoil the diagnosis, but also earn yourself a bad reputation.
You need to donate blood on an empty stomach, because while your stomach is empty, only leukocytes, red blood cells, platelets, protein and antibodies will be visible in the blood. Thus, the analysis will show a more accurate answer.
It is also worth donating biological material (blood) so that specialists from the medical field can easily establish the correct diagnosis, see how antibodies interact with antigens, and ultimately prescribe correct and effective treatment.
For smokers, it is also worth remembering that 2 days before donating blood, you need to refrain from tobacco, as it is also visible in the blood.
The mental and emotional state is also very important, since this is also a factor that affects a person’s well-being and condition. In stressful situations, the blood test will most likely not be unsatisfactory.

Norm and deviations in analysis

When medical professionals conduct such tests, the result is very important, since it can show how the body reacts to the serum.

If a person has no antibodies in his blood at all, then this is a very good result that he does not have any disease. But this happens extremely rarely, since our body is a mechanism that constantly fights with something, sometimes with germs, sometimes with viruses, because every day we walk the streets where there are already germs and bacteria in the air.

Typically, such a blood test is carried out 2 times in order to convince oneself of a particular diagnosis.
Under circumstances where a large number of antibodies are detected in a person, this means that a virus or infection is present in the person’s body. In such circumstances, treatment and examination are prescribed immediately.

How to decipher a blood test

A person who has received a certificate about the result of such an analysis may simply not understand anything, since there will be quite a lot of terminology, which may subsequently be incomprehensible to the average person, but the attending physician who sees such conclusions and results will be able to accurately say the result and interpretation of the analysis .

If, however, something is wrong, the doctor must inform this patient, since this is his job.

As mentioned above, such an analysis is mainly carried out to determine the amount of antibodies; in the normalized version, their amount should be equal to zero.

Bottom line

Most often, people think that donating blood is very scary and painful, but it is not so. Still, we all need to donate blood so that a person knows what is in his blood and whether there are any diseases in his body. It is important to remember that any deterioration in your emotional state, stress or fear of something can cause a storm of emotions in you, which will then be directly reflected in the analysis itself. The main thing is not to worry; after all, this is done for health.

Serology is a branch of immunology that studies the reactions of antigens to serum antibodies.

Serological testing is a technique for studying specific antibodies or antigens in the blood serum of patients. They are based on immune responses. These studies are widely used in the process of diagnosing various infectious diseases and in identifying a person.

Who is prescribed a serological test?

A serological test is prescribed for patients suspected of having any infectious disease. In conflicting situations with diagnosis, this analysis will help to identify the causative agent of the disease. Also, further treatment largely depends on the results of serological studies, since the identification of a specific microorganism contributes to the prescription of specific treatment.

What material is being studied?

Serological studies involve the collection of biological material from the patient in the form of:

Blood serum;

Fecal matter.

The material must be in the laboratory as soon as possible. Otherwise, it can be stored in the refrigerator at +4 or by adding a preservative.

Taking samples

There is no need to specially prepare the patient for taking these tests. The study is safe. A blood test is taken in the morning on an empty stomach, both from the ulnar vein and from the ring finger. After collection, the blood should be placed in a sterile, sealed tube.

Serological blood test

Human blood performs many functions in the body and has a very wide field of activity, therefore there are also many options for blood testing. One of them is serological blood tests. This is a basic analysis carried out to identify certain microbes, viruses and infections, as well as the stage of development of the infectious process. Serological blood tests are used for:

Determining the amount of antibodies against viruses and microbes in the body. To do this, the antigen of the pathogen is added to the blood serum, after which the chemical reaction that occurs is assessed;

Antigen determination by administration;

Blood group definitions.

Serological blood tests are always prescribed twice - to determine the dynamics of the disease. A single determination of the interaction of antigens and antibodies indicates only the fact of infection. To reflect the full picture, where an increase in the number of connections between immunoglobulins and antigens can be observed, a repeat study is necessary.

Serological studies: tests and their interpretation

An increase in the number of antigen-antibody complexes in the body indicates the presence of infection in the patient's body. Carrying out specific chemical reactions with an increase in these indicators in the blood helps determine the disease and its stage.

If the result of the analysis shows the absence of antibodies to pathogens, then this indicates the absence of infection of the body. However, this rarely happens, since the appointment of a serological test already indicates the detection of symptoms of a particular infection.

What can affect the analysis result?

The conditions in which blood is drawn should be carefully monitored. You should not allow anything foreign to get into your bloodstream. The day before the test, you should not overload your body with fatty foods, alcohol and sugary drinks. Stressful situations should be eliminated and physical activity reduced. Biological material must reach the laboratory as soon as possible, since long-term storage of serum leads to partial inactivity of antibodies.

Serological research methods

In laboratory practice, serological blood testing is additional to the main methods presented:

1. Fluorescence reaction, which is carried out in two stages. First, antibodies are detected in the circulating antigen complex. Antiserum is then applied to the control sample, followed by incubation of the preparations. RIF is used to quickly detect the causative agent of the disease in the material being studied. The results of the reactions are assessed using a fluorescent microscope. The nature of the glow, shape, and size of objects are assessed.

2. The agglutination reaction, which is a simple reaction of gluing together discrete antigens with the help of antibodies. Highlight:

Direct reactions used to detect antibodies in the patient's blood serum. A certain amount of killed microbes is added to the serum and causes the formation of a precipitate in the form of flocs. Serological tests for typhoid fever involve a direct agglutination reaction;

Passive hemagglutonation reactions, based on the ability of red blood cells to adsorb antigen on their surface and cause gluing upon its contact with the antibody, and the formation of a visible precipitate. It is used in the process of diagnosing infectious diseases to identify hypersensitivity to certain drugs. When evaluating the results, the appearance of the sediment is taken into account. A ring-shaped sediment at the bottom of the test tube indicates a negative reaction. A lacy sediment with uneven edges indicates the presence of some kind of infection.

3. which is based on the principle of attaching an enzyme tag to antibodies. This allows you to see the result of the reaction by the appearance of enzyme activity or by a change in its level. This research method has several advantages:

Very sensitive;

The reagents used are universal, and they are stable for six months;

The process of recording the analysis results is automated.

The above listed serological research methods have some advantages over the bacteriological method. These methods make it possible to determine the antigens of pathogens in a few minutes or hours. Moreover, these studies can detect pathogen antigens even after treatment and the death of the bacteria that cause it.

Diagnostic value of the study

The results of serological tests are a valuable diagnostic method, but have an auxiliary value. Clinical data still remains the basis for diagnosis. Serological tests are done to confirm the diagnosis if the reactions do not contradict the clinical picture. Weakly positive studies without a clinical picture confirming it cannot become the basis for making a diagnosis. Such results are worth considering when the patient has a history of a similar disease and has received appropriate treatment.

Determining hereditary blood characteristics, confirming or refuting paternity, studying hereditary and autoimmune diseases, establishing the nature and source of infection during epidemics - all this helps to identify serological blood tests. Decoding the results provides information about the presence of specific proteins for infections such as syphilis, hepatitis, HIV, toxoplasmosis, rubella, measles, and typhoid fever.

SEROLOGICAL STUDIES(Latin serum serum + Greek logos doctrine) - methods of immunology that study the specific properties of human or animal blood in order to identify antigens or antibodies using serological reactions.

Beginning of S. and. laid down at the end of the last century, after it was established that the combination of an antigen with an antibody (see Antigen - antibody reaction) is accompanied by a number of phenomena accessible to visual observation - agglutination (see), precipitation (see) or lysis. There is now the possibility of specific recognition of antigens (see) or antibodies (see), if one of these components is known.

In 1897, F. Vidal reported that the blood serum of patients with typhoid fever selectively agglutinates typhoid bacteria and therefore this reaction (see Vidal reaction) can be used in the laboratory. diagnosis of typhoid fever. In the same year, it was shown that filtrates of cultures of plague, typhoid and cholera bacteria, when combined with the corresponding immune sera, form flakes, or precipitates.

The precipitation reaction turned out to be suitable for the detection of any protein antigens. In 1900-1901 K. Landsteiner found that in human erythrocytes there are two different antigens (A and B), and in blood serum there are two agglutinins (a and P), which contributed to the use of the hemagglutination reaction to determine blood groups (see).

The agglutination test to determine blood group and Rh factor is used in obstetric practice, for blood transfusions and tissue transplantation. Antibodies against the Rh factor (see) are incomplete antibodies; they are not capable of direct reaction with Rh-positive erythrocytes, therefore, to detect them, the Coombs reaction is used (see Coombs reaction), based on the detection of incomplete antibodies using antiglobulin sera. The test blood serum is added to red blood cells of known specificity, followed by antiglobulin serum against IgG (indirect Coombs reaction). Fab fragments of incomplete antibodies of the blood serum under study attach to erythrocytes, and anti-IgG antibodies attach to the free Fc fragments of these antibodies, and agglutination of erythrocytes occurs. To diagnose hemolytic anemia, the direct Coombs reaction is used. In the body of such patients, red blood cells combine with antibodies circulating in the blood against the Rh factor. To identify them, anti-IgG antibodies are added to red blood cells taken from the patient. The appearance of red blood cell agglutination confirms the diagnosis of the disease.

The hemagglutination inhibition reaction - HRI (see Hemagglutination) - is based on the phenomenon of prevention (inhibition) by immune serum of hemagglutination of erythrocytes by viruses. The phenomenon of viral hemagglutination is not serol. reaction and occurs as a result of the combination of the virus with red blood cell receptors, however, HAI is a serological reaction used to detect and titrate antiviral antibodies. RTGA is the main method for serodiagnosis of influenza, measles, rubella, mumps, tick-borne encephalitis and other viral infections, the causative agents of which have hemagglutinating properties.

The reaction of passive, or indirect, hemagglutination. It uses red blood cells or neutral synthetic materials (for example, latex particles), on the surface of which antigens (bacterial, viral, tissue) or antibodies are sorbed (see Boyden reaction). Their agglutination occurs with the addition of appropriate sera or antigens. Red blood cells sensitized with antigens are called antigenic erythrocyte diagnosticum and are used to detect and titrate antibodies. Red blood cells sensitized with antibodies are called immunoglobulin erythrocyte diagnosticums (see) and are used to detect antigens:

The passive hemagglutination reaction is used to diagnose diseases caused by bacteria (typhoid and paratyphoid fever, dysentery, brucellosis, plague, cholera, etc.), protozoa (malaria) and viruses (influenza, adenoviral infections, tick-borne encephalitis, Crimean hemorrhagic fever, etc.) . The sensitivity of the passive hemagglutination reaction is not inferior to the method of virus isolation for arenoviral diseases (see), in particular for lymphocytic choriomeningitis. The viral antigen of lymphocytic choriomeningitis is detected in virus carriers (house mice) in a passive hemagglutination reaction with suspensions of extracted organs diluted tens of thousands of times. In case of salmonellosis, the passive hemagglutination reaction detects bacteria at a concentration of up to several hundred microbial bodies in 1 g of feces; dysentery bacteria in food products are detected when there are at least 500 microbial bodies in 1 g of material.

The passive hemagglutination reaction is used in the diagnosis and prevention of viral hepatitis B. In the Soviet Union, to detect the HBs antigen (see Australian antigen) in the blood of patients with acute hepatitis B, a diagnosticum is produced, which is chicken erythrocytes sensitized with goat immunoglobulin against the HBs antigen. A drop of diagnosticum is combined with an equal volume of blood serum from the people being examined, and if HBs antigen is present in it, agglutination occurs. The reaction is capable of capturing up to 1.5 ng/ml of HBs antigen. To detect HBs antibodies, red blood cells with the HBs antigen adsorbed on them, isolated from the blood of patients, are used. The passive hemagglutination reaction is also used to identify the patient’s hypersensitivity to drugs and hormones, for example, penicillin or insulin. In this case, human blood group 0 red blood cells are sensitized with a drug and then used to detect agglutinins to it in the patient’s blood serum.

The passive hemagglutination reaction is used to detect gonadotropic hormone in urine in order to establish pregnancy (see Chorionic gonadotropin). To do this, standard serum for this hormone is incubated with the urine being tested. With the subsequent addition of red blood cells with the hormone sorbed on them, agglutination does not occur (positive response), since the hormone contained in the urine neutralized the agglutinating antibodies.

Reactions based on the phenomenon of precipitation

They are used to determine a wide variety of antigens and antibodies. The simplest example of a qualitative reaction is the formation of an opaque precipitation band at the boundary of the layering of antigen on antibody in a test tube. Various types of precipitation reactions in semi-liquid agar or agarose gels are widely used (double immunodiffusion method according to Ouchterlohn, radial immunodiffusion method, immunoelectrophoresis), which are both qualitative and quantitative in nature (see Immunodiffusion, Immunoelectrophoresis).

To perform double immunodiffusion, a layer of melted gel is poured onto a glass plate and, after hardening, wells with a diameter of 1.5-3 mm are cut out. The test antigens are placed in wells located in a circle, and immune serum of known specificity is placed in the central well. Diffusing towards each other, homologous sera and antigens form a precipitate. With radial immunodiffusion (using the Mancini method), immune serum is added to agar. The antigen placed in the wells diffuses through the agar, and as a result of precipitation with immune serum, opaque rings are formed around the wells, the outer diameter of which is proportional to the concentration of the antigen. A modification of this reaction is used in the diagnosis of influenza to recognize IgM and IgG antibodies (see Immunoglobulins). Influenza antigen is added to the agar, and blood serum is added to the wells. The plates are then treated with immune sera against IgM or IgG antibodies, which helps to detect the reaction of the corresponding antibodies with antigens. The method allows you to simultaneously determine antibody titers and their belonging to a specific class of immunoglobulins.

A type of immunoelectrophoresis is radioimmunophoresis. In this case, after electrophoretic separation of antigens, first immune serum labeled with radioactive iodine against the antigens being determined is poured into a groove cut parallel to the movement of the antigens in the gel, and then immune serum against IgG antibodies is poured, the edges of which precipitate the resulting complexes of the antibody with the antigen. All unbound reagents are washed away, and the antigen-antibody complex is detected by autoradiography (see).

Reactions involving complement. Reactions involving complement (see) are based on the ability of the complement subcomponent Cl(Clq) and then other complement components to attach to immune complexes.

The complement fixation reaction allows antigens or antibodies to be titrated according to the degree of complement fixation by the antigen-antibody complex. This reaction consists of two phases: the interaction of the antigen with the test blood serum (test system) and the interaction of hemolytic serum with sheep red blood cells (indicator system). If the reaction is positive, complement fixation occurs in the test system, and then when erythrocytes sensitized with antibodies are added, hemolysis is not observed (see Complement fixation reaction). The reaction is widely used for the serodiagnosis of visceral syphilis (see Wasserman reaction) and viral infections (see Virological studies).

Cytolysis. Antibodies against cellular structures can, with the participation of complement, dissolve the cells bearing these structures. Lysis of red blood cells is easily assessed by the degree and intensity of hemoglobin release. Nuclear cell lysis is assessed by calculating the percentage of dead cells that are not stained with methylene blue. Radioactive chromium is also often used, which is previously chemically bonded to the cells. The number of destroyed cells is determined by the amount of unbound chromium released during cell lysis.

The reaction of radial hemolysis of erythrocytes can occur in the gel. A suspension of sheep red blood cells is placed in an agarose gel, adding complement; Wells are made in the layer frozen on the glass and hemolytic serum is added to them. A hemolysis zone will form around the wells as a result of radial diffusion of antibodies. The radius of the hemolysis zone is directly proportional to the serum titer. If you adsorb any antigen on erythrocytes, for example, the glycoprotein hemagglutinin of influenza virus, rubella or tick-borne encephalitis, then you can reproduce the phenomenon of hemolysis with immune sera to these viruses. The radial hemolysis reaction in a gel has found application in the diagnosis of viral infections due to its ease of production, insensitivity to serum inhibitors, and the ability to titrate blood serum according to the diameter of the hemolysis zone without resorting to serial dilutions.

Immune adhesion. Red blood cells, platelets and other blood cells have receptors for the third component of complement (C3) on their surface. If the appropriate immune serum and complement are added to an antigen (bacteria, viruses, etc.), an antigen-antibody complex coated with the C3 component of complement is formed. When mixed with platelets, due to the C3 component of complement, the antigen-antibody complex will settle on the cells and cause their agglutination (see Immune adhesion). This reaction is used to determine antigens of the HLA system (see Transplantation immunity) and in the study of a number of viral infections (tick-borne encephalitis, dengue fever), which are accompanied by immunopathol. processes and circulation in the blood of viral antigens in combination with antibodies.

The neutralization reaction is based on the ability of antibodies to neutralize certain specific functions of macromolecular or soluble antigens, for example, enzyme activity, bacterial toxins, and pathogenicity of viruses. In bacteriology, this reaction is used to detect antistreptolysins, antistreptokinase and antistaphylolysins. The neutralization reaction of toxins can be assessed by biol. effect, for example, antitetanus and antibotulinum serums are titrated (see Toxin - antitoxin reaction). A mixture of toxin and antiserum administered to animals prevents their death. Various versions of the neutralization reaction are used in virology. By mixing viruses with the appropriate antiserum and injecting this mixture into animals or cell cultures, the pathogenicity of the viruses is neutralized.

Reactions using chemical and physical labels

Immunofluorescence, developed by A. N. Coons in 1942, is used for serol. reactions of fluorochrome-labeled sera (see Immunofluorescence). Fluorochrome-labeled serum forms an antigen-antibody complex with the antigen, which becomes accessible to observation under a microscope in ultraviolet rays, which excite the fluorochrome. The direct immunofluorescence reaction is used to study cellular antigens, detect virus in infected cells, and detect bacteria and rickettsiae in smears. Thus, to diagnose rabies, prints of pieces of the brain of animals suspected of carrying the virus are treated with luminescent anti-rabies serum. If the result is positive, lumps of bright green color are observed in the protoplasm of nerve cells. Express diagnostics of influenza, parainfluenza and adenoviral infection is based on the detection of viral antigens in fingerprint cells from the nasal mucosa.

The more widely used method is indirect immunofluorescence, based on the detection of the antigen-antibody complex using luminescent immune serum against IgG antibodies and used to detect not only antigens, but also titration of antibodies. The method has found application in the serodiagnosis of herpes, cytomegalips, and Lassa fever. In the laboratory, a stock of preparations of antigen-containing cells, for example, VERO cells grown on pieces of thin glass and infected with a virus or chicken fibroblasts fixed with acetone should be stored at -20°. The test blood serum is layered onto the preparations, the preparation is placed in a thermostat at f 37° to form immune complexes, and then, after washing off unbound reagents, these complexes are detected with labeled luminescent serum against human globulins. By using labeled immune sera against IgM or IgG antibodies, it is possible to differentiate the type of antibodies and detect the early immune response by the presence of IgM antibodies.

In the enzyme-immunological method, antibodies conjugated with enzymes are used, ch. arr. horseradish peroxidase or alkaline phosphatase. To detect the combination of labeled serum with antigen, a substrate is added that is decomposed by the enzyme attached to the serum, producing a yellow-brown (peroxidase) or yellow-green (phosphatase) color. Enzymes are also used that decompose not only chromogenic, but also lumogenic substrates. In this case, with a positive reaction, a glow appears. Like immunofluorescence, the enzymatic immunological method is used to detect antigens in cells or titrate antibodies on antigen-containing cells.

The most popular type of enzyme-immunological method is immunosorption. On a solid carrier, which can be cellulose, polyacrylamide, dextran and various plastics, the antigen is absorbed. Most often, the surface of the micropanel wells serves as the carrier. The test blood serum is added to the wells with the sorbed antigen, then the enzyme-labeled antiserum and substrate. Positive results are taken into account by changes in the color of the liquid medium. To detect antigens, antibodies are sorbed onto the carrier, then the test material is added to the wells and the reaction is carried out with an enzyme-labeled antimicrobial serum.

The radioimmunological method is based on the use of radioisotope labels of antigens or antibodies. It was originally developed as a specific method for measuring the levels of hormones circulating in the blood. The test system was an isotope-labeled hormone (antigen) and antiserum to it. If a material containing the desired hormone is added to such an antiserum, it will bind part of the antibodies; upon subsequent addition of a labeled titrated hormone, a reduced amount of it will bind to the antibodies compared to the control. The result is assessed by comparing the curves of the bound and unbound radioactive tracer. This type of method is called a competitive reaction. There are other modifications of the radioimmunological method. The radioimmunological method is the most sensitive method for determining antigens and antibodies, used for the determination of hormones, drugs and antibiotics, for the diagnosis of bacterial, viral, rickettsial, protozoal diseases, for the study of blood proteins, tissue antigens.

Comparative characteristics and use of serological research methods in medical practice

Methods S. and. are constantly being improved to increase sensitivity and versatility of use. Initially serol. diagnosis was based on the detection of antibodies. With the advent of the mid-20th century. immunofluorescence and passive hemagglutination reactions, which are more sensitive, made it possible to detect not only antibodies, but also antigen directly in material from patients. Enzyme-immunological and radioimmunological methods, which are 2-3 orders of magnitude more sensitive than immunofluorescence and passive hemagglutination, are close to biological methods. detection of bacteria and viruses. The scope of their application for the detection of both antigens and antibodies is theoretically unlimited.

Serodiagnosis info. diseases is based on the appearance of antibodies to an isolated or suspected pathogen, regardless of whether the pathogen was detected in the acute stage of the disease. Pairs of blood serum taken at the onset of the disease and 2-3 weeks later are examined. later. An increase in antibodies in the second blood serum of at least 4 times compared to the first is diagnostically significant. It also matters which class of immunoglobulins the antibodies are represented by. IgM antibodies are detected at the end of the acute period of the disease and in the early stage of convalescence. IgG antibodies appear later in convalescence and circulate for a long time. If IgM antibodies to the rubella virus are detected in a woman in the first trimester of pregnancy, this serves as a basis for terminating the pregnancy, since during this period the fetus is especially sensitive to the virus. With different inf. diseases, the most specific and convenient methods are selectively used.

S. and. widely used in epidemiology. Systematic collection and examination of blood samples from various population groups makes it possible to understand the contacts of the population with the source of infectious agents. diseases. Studying the level of collective immunity allows us to identify high-risk groups and plan vaccination activities, and study the geographic spread of infections. S. and. different age groups of the population made it possible, for example, to retrospectively identify the circulation of different variants of the influenza virus during certain periods of time.

S. and. are of great importance in the study of hereditary diseases (see) and autoimmune diseases, accompanied by the appearance of tissue- and organ-specific antibodies that destroy the corresponding target cells, as well as in oncology for the detection of tumor antigens. Thus, the immunodiagnosis of liver cancer is based on the determination of alpha-fetoprotein and other embryonic antigens in the blood serum of patients using the immunodiffusion method and the radioimmunological method.

Significant scientific progress in the study of the fine antigenic structure of cellular antigens, antigens of bacteria and viruses is achieved through the use of serol. reactions of monoclonal antibodies, which can be obtained to individual antigen determinants.

Bibliography: Research methods in immunology, ed. I. Lefkovits and B. Pernis, trans. from English, M., 1981; Guide to immunology, ed. O. E. Vyazova and Sh. X. Khodzhaeva, M., 1973; Guide to Clinical Laboratory Diagnostics, ed. V.V. Menshikova, M., 1982; Immunology, ed. by J.-F. Bach, N. Y., 1978.

S. Ya. Gaidamovich.