The agglutination reaction is based on the specific interaction of antibodies (agglutinins) with whole microbial or other cells. As a result of this interaction, agglomerate particles are formed that precipitate (agglutinate). Bacteria, protozoa, fungi, yeast, rickettsia, erythrocytes and other cells, both living and killed, can participate in the agglutination reaction. The reaction occurs in two phases: the first is a specific combination of antigen and antibody, the second is non-specific, i.e. the formation of a visible agglutinate. Precipitation of agglutinate occurs in the presence of electrolytes, such as sodium chloride. The microorganisms in the agglutinate remain alive, but lose their mobility.
The agglutination reaction is widely used for serological diagnosis of infectious diseases and determination of the antigenic structure of isolated microbes. To determine the antigenic structure of a pathogen isolated from the body of a patient or a carrier, specific immune serum is used, obtained by immunizing animals (rabbit, donkey, sheep) with certain microorganisms. Identification of the microbe is carried out in an agglutination reaction on glass with adsorbed or monoreceptor sera or in test tubes with specific agglutinating sera. Adsorbed sera contain antibodies only to antigens specific to a given microbe, and monoreceptor sera contain antibodies only to one specific antigen of the pathogen.
Species sera contain antibodies to all antigens of a particular microbe.
Whether the isolated microorganism culture belongs to this species determined by agglutination with a known serum to the antibody titer indicated on the label of the serum ampoule. The antibody titer of a serum is considered to be its last dilution, in which agglutination of the culture of microbes used to immunize the animal is still observed. Adsorbed and monoreceptor sera are usually used undiluted in the glass agglutination reaction.
When determining the presence of antibodies in the patient’s blood serum, it is diluted with an isotonic sodium chloride solution starting from a dilution of 1: 50 to 1: 800 or more. A suspension of live or killed microbes is added to each dilution. Preparations containing microbes killed by heat or formaldehyde are called diagnosticums. Diagnosticums obtained by heating microorganism cultures contain only somatic antigens. When using only formaldehyde, microbes retain their flagellar antigens.
In the presence of antibodies in the patient’s blood, the diagnostic test taken in the reaction sticks together and a precipitate (agglutinate) forms on two test tubes. In this case, the results of the agglutination reaction are regarded as positive. In the control tube, into which isotonic sodium chloride solution and diagnosticum are added, the suspension of microbes should be homogeneous (negative agglutination reaction).
The results of the agglutination reaction in some diseases, such as leptospirosis, are taken into account only microscopically in a dark field of view of a microscope (microagglutination). To make a serological diagnosis of a disease, take into account diagnostic disease. It usually corresponds to a serum dilution of 1:100 or 1:200.
Antibodies in the patient’s blood serum can be detected using the agglutination reaction in cases of typhoid fever and paratyphoid fever (Vidal reaction), brucellosis (Wright reaction), tularemia, etc.
Castellani's reaction. For some infectious diseases or immunization with microorganisms containing group antigens, in the blood serum, in addition to antibodies specific to a given type, group antibodies also appear. In this case, related bacterial species will be agglutinated by the resulting sera.
Castellani proposed a method for the adsorption of group antibodies from immune sera, based on their removal with the help of microorganisms of related species that have group antigens but lack specific ones. A culture of such microorganisms added to the serum adsorbs nonspecific group antibodies, and after removal of the antigen-antibody complex by centrifugation, only specific immunoglobulins remain in the serum. Sera processed according to the Castellani method can be used in the agglutination reaction as highly specific.
Agglutination is the gluing of bacteria as a result of the interaction of specific ATs with them. To carry out RA, three components are required: 1) AG (agglutinogen); 2) AT (agglutinin); 3) electrolyte solution (isotonic sodium chloride solution). Only corpuscular antigens (bacteria, red blood cells, antigen-loaded latex particles) take part in the agglutination reaction.
Agglutination reaction on glass. Place a drop onto a grease-free glass slide using a pipette. diagnostic serum(serum dilution 1:10 - 1:20). Using a bacteriological loop, a pure culture of the microorganism under study is taken from the surface of the slanted agar, transferred to a drop of serum and mixed. The result of the reaction is taken into account with the naked eye after 3-5 minutes. If the reaction is positive, the appearance of flakes (large or small) is noted in a drop of serum, clearly visible against a dark background when the slide is shaken. In case of a negative reaction, the liquid remains uniformly cloudy.
Agglutination reaction in test tubes. 1 ml of physiological solution is added to a row of test tubes. An equal volume of the blood serum being tested is added to the first test tube. Serial two-fold dilutions of serum are prepared (serum titration), after which 2 drops of a suspension of inactivated bacteria (diagnosticum) are added to each test tube. The tubes are placed in a thermostat at 37 °C for 2 hours. The reaction proceeds with the formation of small flakes, invisible to the naked eye, so the results are recorded under slight magnification in a special device - an agglutinoscope. The intensity of agglutination is taken into account according to the “four plus” system: complete agglutination - 4+, partial agglutination - 3+ or 2+, questionable result - +. The last dilution in which agglutination of 2+ is observed is taken as the antibody titer in the test serum.
An agglutination reaction in test tubes (an extensive agglutination reaction) is carried out to determine the titer of antibodies to the causative agents of typhoid and paratyphoid fever (Vidal reaction), brucellosis (Wright reaction), and typhus (Weigl reaction).
Agglutination (from Latin agglutinatio - gluing) is the gluing (connection) of antigen-bearing corpuscular particles (whole cells, latex particles, etc.) with molecules of specific antibodies in the presence of electrolytes, which ends in the formation of flakes or sediment (agglutinate) visible to the naked eye. The nature of the sediment depends on the nature of the antigen: flagellated bacteria produce a coarse flocculent sediment, flagellated and noncapsular bacteria produce a fine-grained sediment, and capsular bacteria produce a stringy sediment. A distinction is made between direct agglutination, in which the own antigens of a bacterial or any other cell, such as erythrocytes, directly participate in the interaction with specific antibodies; and indirect, or passive, in which bacterial cells or erythrocytes, or latex particles are carriers not of their own, but of foreign antigens (or antibodies) sorbed on them to identify antibodies (or antigens) specific to them. The agglutination reaction mainly involves antibodies belonging to the IgG and IgM classes. It occurs in two phases: first, a specific interaction of the active center of antibodies with the determinant of the antigen occurs; this stage can occur in the absence of electrolytes and is not accompanied visible changes reacting system. For the second stage - the formation of agglutinate - the presence of electrolytes is necessary, which reduce electric charge antigen + antibody complexes and accelerate the process of their gluing. This phase ends with the formation of an agglutinate.
Agglutination reactions are carried out either on glass or smooth cardboard plates, or in sterile agglutination tubes. Agglutination reactions (direct and passive) on glass are usually used as accelerated method detection of specific antibodies in the patient’s serum (for example, with brucellosis) or for serological identification of the pathogen. In the latter case, well-purified (adsorbed) diagnostic sera containing only monoreceptor antibodies or a set of them to various antigens are usually used. The undoubted advantage of the agglutination reaction on glass is the simplicity of its implementation and the fact that it takes several minutes or even seconds, since both components are used in concentrated form. However, it has only a qualitative value and is less sensitive than a test tube. An extensive agglutination reaction in test tubes gives more accurate results, because it allows you to determine the quantitative content of antibodies in the serum (establish its titer) and, if necessary, register the fact of an increase in antibody titer, which has diagnostic value. To set up the reaction, a serum diluted in a certain way with a 0.85% NaCl solution and an equal volume (usually 0.5 ml) of a suspension of a standard diagnosticum (or test culture) containing 1 billion bacteria in 1 ml are added to agglutination tubes. The results of the agglutination reaction are recorded first after 2 hours of incubation of the tubes at 37 °C and finally after 20-24 hours according to two criteria: the presence and size of the precipitate and the degree of transparency of the supernatant liquid. The assessment is carried out according to the four-cross system. The reaction is necessarily accompanied by serum and antigen control. In cases where a detailed agglutination reaction in a test tube is performed for serological identification of the pathogen, it has diagnostic value if the reaction is assessed as positive when the diagnostic serum is diluted to at least half its titer.
It must be taken into account that when mixing solutions of homologous antigens and antibodies, visible manifestations of the agglutination reaction are not always observed. A precipitate forms only at certain optimal ratios of both reaction components. Outside these limits, with a significant excess of antigen or antibodies, no reaction is observed. This phenomenon is called the “prozone phenomenon.” It is observed both in the agglutination reaction and in the precipitation reaction. The appearance of the prozone in immune reactions is explained by the fact that the antigens involved in them, as a rule, are polydeterminant, and the molecules IgG antibodies have two active centers. With an excess of antibodies, the surface of each antigen particle is covered with antibody molecules so that there are no free determinant groups left, so the second, unbound active center of the antibodies cannot interact with another antigen particle and bind them to each other. The formation of a visible agglutinate or precipitate is also suppressed when there is an excess of antigen, when not a single free active center of the antibodies remains, and therefore the antigen + antibody + antigen complexes can no longer enlarge.
Options for accelerated agglutination reactions. Reaction passive hemagglutination and its variants
The classic agglutination reaction involves the use of corpuscular antigens. However, soluble antigens may also be involved. To make this possible, such antigens are adsorbed onto immunologically inert particles. Particles of latex or bentonite can be used as a carrier, but currently animal or human erythrocytes are most often used, improving their adsorbing properties by treating them with solutions of tannin, formalin or benzidine. Red blood cells that have adsorbed an antigen on themselves are called sensitized by this antigen, and immune reaction in which they participate is an indirect or passive hemagglutination reaction (IRHA, or RPHA), since red blood cells participate in it passively.
RPGA is placed in special polystyrene plates with holes having a hemispherical bottom. When using it for serological diagnostics In these wells, two-fold dilutions of the test serum are prepared in physiological solution, and then a suspension of sensitized erythrocytes is added to it as a diagnostic agent. The results are recorded after 2 hours of incubation at 37 °C using the four-cross system. With a positive reaction, agglutinated red blood cells settle to the bottom of the hole and evenly cover it in the form of an inverted umbrella. At negative reaction red blood cells also settle, the liquid becomes transparent, the sediment looks like a small “disc” in the center of the hole. The serum titer in RPHA is considered to be its last dilution, which still gives pronounced hemagglutination without significant signs of the presence of a “disc”.
RPGA can also be used as an accelerated method of bacteriological diagnostics to detect directly in the test material unknown bacteria, viruses, toxins, for example, plague pathogens, staphylococcal enterotoxins, etc. With this version of RPGA, erythrocytes that have adsorbed known for their specificity are used as a known component of the reaction antibodies - antibody erythrocyte diagnosticum. If the test material contains a sufficient amount of a known antigen, RPGA will be positive.
Options for using RPHA are: antigen neutralization reaction (RNAg), antibody neutralization reaction (RNAb), passive hemagglutination inhibition reaction (PHA). For these reactions, antigen and antibody erythrocyte diagnostics are used. Two mutually controlling unidirectional reactions can be used simultaneously, for example, RPHA with an antigen diagnosticum and RNAg with an antibody erythrocyte diagnosticum.
The antibody neutralization reaction (RNAb) consists of mixing a suspension containing the desired antigen with a specific immune serum containing known antibodies in appropriate volumes and incubating at 37 °C for two hours. After this, an antigenic erythrocyte diagnosticum is added. The mixture is shaken and left at room temperature. The results are taken into account after 3-4 hours and finally after 18-24 hours. If the test material contains antigen, it will bind antibodies (neutralize them), and therefore hemagglutination will not occur.
The antigen neutralization reaction (RNAg) is performed using the same principle. Only in this case are antibodies detected in the test material. A specific antigen added to such a test material will bind to the antibodies contained in it, i.e., neutralization of the antigen by antibodies will occur, and therefore hemagglutination will not occur when adding an antibody erythrocyte diagnosticum.
Coagglutination reaction. It is one of the options for a passive, i.e., accelerated agglutination reaction on glass mediated by antibody-carrying cells. This reaction is based on a unique property Staphylococcus aureus, which contains protein A in its cell wall, binds to the Fc fragments of IgG and IgM. In this case, the active centers of antibodies remain free and can interact with specific determinants of antigens. A drop of a 2% suspension of staphylococci, sensitized with appropriate antibodies, is applied to a glass slide, and a drop of a suspension of the bacteria being studied is added. If the antigen matches the antibodies, a clear agglutination of the staphylococci loaded with antibodies occurs within 30-60 s.
Latex agglutination reaction (LAR). The carrier of antibodies in this diagnostic system are small standard latex particles. The reaction is performed using the micromethod in wells on glass. The main condition for the successful staging of PAH is strict adherence to the quantitative ratios of the system components: 10 μl of a latex preparation sensitized with antibodies is added to 50 μl of the test material. The specificity of PAH is controlled using three control tests contained in commercial test systems: known positive reaction, obviously negative reaction and quality control of latex suspension for PAH-unsensitized (not carrying antibodies) latexes with the test material. In our country, polystyrene monodisperse latexes with different particle diameters (0.3; 0.66; 0.75; 0.8 μm) are used as carriers of specific antibodies. LAG is used for the rapid detection of microorganisms or their antigens in the test material.
Immunomagnetic detection of antigens. One of the options for accelerated agglutination reaction on glass is associated with the use of supermagnetic polymer particles coated with specific antibodies. One such particle binds up to 107-108 cells of microorganisms, due to which sensitivity this method reaches 5 CFU/ml. Immunomagnetic detection of microorganisms can be used in combination with CPR.
Aggregate hemagglutination reaction (AHA). Allows you to quickly detect in the blood of patients both freely circulating antigens (antigenemia) and antigens associated with antibodies - circulating immune complexes (CIC). For RAHA, red blood cells sensitized with appropriate antibodies are used. The addition of the patient's blood serum, which contains antigens, to sensitized erythrocytes on which antibodies are fixed, leads to gluing (agglutination) of erythrocytes and immune complexes.
Antiglobulin Coombs test (R. Coombs reaction). Full (divalent) antibodies are determined using direct and passive agglutination reactions. Incomplete (monovalent, blocking) antibodies are not detected by these methods, since, when they combine with the antigen, they block it, but cannot cause aggregation of the antigen into large conglomerates. Incomplete (blocking) antibodies are those in which only one active center functions; the second active center does not work for an unknown reason. To detect incomplete antibodies, a special Coombs reaction is used (Fig. 72). The reaction involves: the patient’s serum, in which incomplete antibodies are determined, corpuscular antigen-diagnosticum, antiglobulin serum containing antibodies to human globulin. The reaction occurs in two stages:
Interaction of antigen with incomplete antibodies. There are no visible manifestations. The first stage is completed by washing the antigen from the remaining serum of the patient.
Interaction of antiglobulin serum obtained as a result of immunization of an animal with human globulin with incomplete antibodies adsorbed on the antigen. Due to the fact that antiglobulin antibodies are bivalent, they bind two monovalent antibodies of separate AG + complexes incomplete antibody, which leads to their gluing and the appearance of visible sediment.
Agglutination is the gluing and precipitation of microbes or other cells under the influence of antibodies in the presence of an electrolyte (isotonic sodium chloride solution). Groups of glued bacteria (cells) are called agglutinate. The following components are required for the agglutination reaction:
1. Antibodies (agglutinins) that are found in the serum of a sick or immune animal.
2. Antigen - a suspension of living or killed microbes, red blood cells or other cells.
3. Isotonic (0.9%) sodium chloride solution.
The agglutination test for serodiagnosis is used for typhoid fever and paratyphoid fever (Vidal reaction), brucellosis (Wright and Heddleson reaction), tularemia, etc. The antibody is the patient's serum, and the antigen is a known microbe. When identifying microbes or other cells, their suspension is used as an antigen, and a known immune serum is used as an antibody. This reaction is widely used for diagnostics intestinal infections, whooping cough, etc.
Methods for staging RA
Approximate RA on glass
Deployed RA
(volume method)
Coagglutination reaction
Unfolded RA on glass (seroidentification)
Agglutination reaction on glass. Two drops of specific (adsorbed) serum and a drop of isotonic sodium chloride solution are applied to a fat-free glass slide. Non-adsorbed serums are pre-diluted in a ratio of 1:5 - 1:100. Drops must be applied to the glass so that there is a distance between them. The culture is thoroughly ground on glass with a loop or pipette, and then added to a drop of isotonic sodium chloride solution and to one of the serum drops, stirring in each until a homogeneous suspension is formed. A drop of serum without culture is a serum control.
Attention! You cannot transfer the culture from the serum to a drop of isotonic sodium chloride solution, which serves as an antigen control. The reaction takes place at room temperature for 1-3 minutes. If the serum control remains transparent, a uniform turbidity is observed in the antigen control, and agglutinate flakes appear in the drop where the culture is mixed with serum against the background of a clear liquid, the reaction result is considered positive.
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Diagnostic Physiological
serum + culture solution + culture
Detailed agglutination reaction (volume method). Serial, most often twofold, dilutions of serum are prepared. The method is called volumetric. To determine the antibody titer in blood serum, take 6 tubes. Pour 1 ml of the original serum dilution 1:50 into the first test tube and add 1 ml of saline solution into all 6 test tubes using a graduated pipette. The first test tube will yield a serum dilution of 1:100 with a volume of 2 ml. Transfer 1 ml from the first test tube to the second test tube, where the dilution becomes 1:200. So make a series of serial dilutions of the serum in the first 5 test tubes (1:100, 1:200, 1:400, 1:800, 1:1600). From the fifth test tube, pour 1 ml into the disinfectant solution. Add 2 drops of diagnosticum to all 6 test tubes. The sixth tube is a culture control, as it contains only saline solution and diagnosticum.
Such control is necessary to exclude spontaneous agglutination of the culture. The tubes are shaken and placed in a thermostat at a temperature of 37°C for 2 hours, and then left for a day at room temperature, after which the results of the agglutination reaction are recorded. When performing an agglutination reaction with the sera of children in the first months of life, due to the functional inferiority of antibody formation, it is necessary to identify lower antibody titers, which is taken into account when diluting the serum. The initial serum dilution is 1:25. In the first test tube, a dilution of 1:50 is obtained, then 1:100, etc.
At positive result reactions in test tubes, sticky cells in the form of grains or flakes are visible against the background of a clear liquid. The agglutinate gradually settles to the bottom in the form of an “umbrella”, and the liquid above the sediment becomes clear. The antigen control is uniformly turbid.
Based on the nature of the sediment, fine- and coarse-grained (flaky) agglutination is distinguished. Fine-grained agglutination is obtained when working with O-sera. Coarse-grained - when motile microbes interact with flagellar H-sera. It occurs faster than fine-grained, and the resulting sediment is very loose and easily broken.
The intensity of the reaction is expressed as follows:
All cells have settled, the liquid in the test tube is completely transparent. The result of the reaction is sharply positive;
There is less sediment, the liquid does not clear completely. The result of the reaction is positive;
There is even less sediment, the liquid is cloudier. The result of the reaction is doubtful;
There is a slight sediment at the bottom of the test tube, the liquid is cloudy. Questionable reaction result;
There is no sediment, the liquid is uniformly cloudy, as in the antigen control. Negative reaction result
Agglutination reaction Agglutination reaction
(RA) - a method of identifying and quantification Ag and At, based on their ability to form agglomerates visible to the naked eye. In the department of infectious diseases. diseases or for other purposes is used to identify unknown microbes and cells, to determine the presence and amount of Ab in blood and other fluids. The determination principle is based on the specificity of the interaction between Ag and Ab and consists in finding the known from the unknown. There are many options for RA: quantitative and qualitative, test tube and glass, volumetric and droplet, conventional, accelerated and express methods. To stage RA you need: 1) s-ka blood. In the variant with determining the type (var) of bacteria, industrial agglutinating tests are used, produced by immunizing rabbits. In the variant with determination of the type of Ab, a blood sample is taken from the test. people or animals. The solution must be sterile and free of suspended particles. Prepare the basic dilution in saline solution. It should be 2-4 times lower than the diagnostic titer for this disease; 2) Ag. In the version of the reaction with determination of the Ab type, industrial diagnostic kits are used; in the variant with the determination of Ag, diagnosticums are prepared themselves in the form of a 1-3 billion suspension in a saline solution of 18-20-hour agar (less often broth) test. microbe inactivated by heating in a water bath at 70°C for 1 hour or by 24-hour incubation at 37°C with formaldehyde (final concentration 0.2%); 3) electrolyte in the form of saline solution. Staging technique volumetric serial tube RA to determine the Ab titer in s-ki: several rows of working dilutions are prepared from the main dilution of s-ki. The number of rows depends on the number of diagnosticums taken into the experiment; the number and dilution factors are determined by the diagnostic titer of the suspected disease. The series must at least contain a dilution corresponding to the diagnostic Ab titer, two dilutions below and two dilutions above it. for example, if the diagnostic titer is 1:100, then with the volumetric method of staging RA the following dilutions should be prepared: 1:25, 1:50, 1:100, 1:200, 1"400; with the drip method, the first dilution (1:25) is not needed, but another higher dilution is needed - 1:800. scientific research s-ku is titrated to a negative reaction. It is diluted as follows: 0.25 ml of saline solution is poured into all test tubes, except the 1st one, when the reaction is carried out in a volume of 0.5 ml, and 0.5 ml when the reaction is carried out in a volume of 1 ml. Pour 0.25 (0.5) ml of the main dilution into the 1st and 2nd test tubes, from the 2nd test tube, into the cut volume and breeding s-k and increased 2 times, 0.25 (0.5) ml is transferred to the 3rd, from the 3rd to the 4th, etc. to the last, from the cut 0.25 (0.5) ml is poured into everything to balance the volumes. Each dilution is carried out using a separate pipette. If several diagnosticums are taken into experiment, then for each of them its own series of dilutions is prepared in the same way. Diagnosticum is added to each dilution of the test tube in a volume equal to the volume of the test tube, as a result of which the dilution in each test tube is doubled. The experiment corresponds to the s-ki control (0.25 - 0.5 ml of the main dilution of s-ki and the same amount of saline solution) and the Ag control (0.25 - 0.5 ml of diagnosticum and the same amount of saline solution). If several diagnosticums are used in the experiment, then each has its own antigen control. The rack with test tubes is shaken well and placed in a thermostat at 37°C for 4 hours, and then left at room temperature until the next day, after which the PA is recorded based on the amount of sediment and the degree of clearing of the liquid. Determination of these indicators, depending on the nature of the agglutinates, is carried out with the naked eye on a dark background, in an agglutinoscope or over the concave surface of a microscope mirror. Accounting begins with controls: control C should be transparent, Ag should be uniformly cloudy (after shaking the tube). If the controls are good, establish the presence and degree of agglutination in all test tubes, which are designated by pluses: large sediment and complete clearing of the liquid - 4 pluses; large sediment and incomplete clearing of the liquid - 3 pluses; noticeable sediment and noticeable clearing of the liquid are 2 pluses. After this, the titer is determined: the highest dilution with an agglutination intensity of at least 2 pluses. Titre research s-ki are compared with the diagnostic titer for this disease. If the titer is examined. s-ki is 2 times lower than the diagnostic value, the reaction is assessed as doubtful; if the titer is equal diagnostic - how weakly positive; if it is 2-4 times higher, it is considered positive; if it is 8 or more times higher, it is considered sharply positive. With the widespread distribution of Ab healthy people To assess RA, an increase in Ab titer is used. To determine the type of Ar in serial RA, the number of rows must correspond to the number taken for identification diagnostic tests. From the main dilution of the diagnostic test, a series of successive two-fold dilutions are prepared in the same way as in RA to determine the Ab titer. Dilution factors depend on the titer of the agglutinating test. In the experiment, the presence of a dilution equal to the titer of the test is necessary, as well as 2, 4, 6, 8 times lower than it. For example, if the titer of the diagnostic test is 1 3200, then you should use dilutions 1 3200, 1 1600, 1 800, 1 400, 1 200 The same volume of tested Ag is added to the dilutions of the test, as a result, the dilution of the test increases by 2 times. 2 controls of test and Ag are added to the experiment. If in the experiment several s-k are involved, then each of them needs its own control. Upon completion of the reaction, the stand is shaken vigorously and placed in a thermostat at 37 ° C. The results are taken into account as described above. Evaluation of the reaction has features. In order to draw a conclusion about the compliance of the study. Ag taken into the experiment, the titer of the reaction must correspond to at least half the titer of the standard diagnostic test. Titers of 1 4 and below are considered as a group reaction Drip md staging of RA differs from volumetric in that s-ku is diluted in a volume of 1 ml, Ag is used in a higher concentration (10 billion/ml) and it is added 1 - 2 drops into a test tube Dilution of the drug after adding Ag is considered unchanged. Otherwise, the method of setting, recording and evaluation is similar to the volumetric method
(Source: Dictionary of Microbiology Terms)
Agglutination reaction (from lat. agglutinatio- gluing) - gluing of corpuscles (bacteria, red blood cells, etc.) by antibodies in the presence of electrolytes.
Agglutination reaction manifests itself in the form of flakes or sediment consisting of corpuscles (for example, bacteria) “glued together” by antibodies (Fig. 7.37). The agglutination reaction is used to: determine the pathogen isolated from the patient; determination of antibodies in the patient’s blood serum; determination of blood groups.
Rice. 7.37 a, b. Agglutination reaction withIgM-antibodies (a) andIgG-antibodies (b)
1. Determination of the pathogen isolated from the patient Approximate reaction agglutination on glass (Fig. 7.38). A suspension of bacteria isolated from the patient is added to a drop of agglutinating serum (1:20 dilution). A flocculent precipitate forms.
Rice. 7.38.
An extensive agglutination reaction with a pathogen isolated from a patient (Fig. 7.39). A suspension of bacteria isolated from the patient is added to the dilutions of the agglutinating serum.
Rice. 52
2. Determination of antibodies in the patient’s blood serum
Detailed agglutination reaction with the patient’s blood serum (Fig. 7.39). Diagnosticum is added to dilutions of the patient's serum.
- Agglutination with O-diagnosticum (bacteria killed by heat, retaining O-antigen) occurs in the form of fine-grained agglutination.
- Agglutination with H-diagnosticum (bacteria killed by formaldehyde, retaining the flagellar H-antigen) is large and occurs faster.
3. Agglutination reaction for determining blood groups The agglutination reaction to determine blood groups is used to establish the ABO system (Table b) using agglutination of erythrocytes with immune serum antibodies against blood group antigens A (I), B (III). The control is: serum that does not contain antibodies, i.e. serum AB (IV) blood group; antigens contained in red blood cells of groups A (II), B (III). The negative control does not contain antigens, i.e., group O (I) erythrocytes are used.
Table 7.6. Determination of ABO blood groups
| Reaction results |
Group |
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belonging |
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researched |
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red blood cells with |
serum (plasma) |
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standard |
with standard |
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serums | ||
