Direct Coombs test analysis. Indirect Coombs test (antiglobulin test, detection of incomplete anti-erythrocyte antibodies), blood

Coombs test– an assay to detect antibodies attached to a surface or dissolved in plasma. It is used to detect immunization and antibodies to red blood cells. The second name is antiglobulin test. It can be direct or indirect.

At direct antiglobulin test detects antibodies fixed on the surface of red blood cells. Carried out if there is a suspicion of, for others autoimmune diseases, after taking medications (methyldopa, penicillin, quinine) and.

Red blood cells have been sensitized in vivo - antibodies are already firmly attached to them, and the addition of antiglobulin serum (anti-IgG) causes the sensitized cells to stick together, which is visible to the naked eye.

Indirect Coombs test detects anti-erythrocyte antibodies in blood plasma, it is performed before blood transfusion and during.

Anti-erythrocyte antibodies are a type of autoantibodies, i.e. antibodies against your own tissues. Autoantibody occurs during pathological reactivity immune system for some drugs, for example high doses penicillin.

Red blood cells on their surface contain various chemical structures(glycolipids, saccharides, glycoproteins and proteins), in medicine called antigens. A person inherits from his parents a specific map of antigens on each red blood cell.

Antigens are combined into groups and then the blood is divided into several groups - according to the AB0, Rh, Kell, Lewis, Kidd, Duffy system. The most famous and significant in the work of a doctor are AB0 and the Rh factor (Rh).

AB0 system

A person's Rh status is determined by the presence of these antigens. A particularly important antigen of erythrocytes is antigen D. If it is present, then they speak of Rh positive blood RhD, and if it’s not there – oh Rh negative Rhd.

If the corresponding antibody attaches to the erythrocyte antigens, the erythrocyte is destroyed - hemolysis.

Indications

The main indication for directantiglobulin test- suspicion of hemolytic anemia. It is most often performed for primary autoimmune diseases. hemolytic anemia, hemolysis in rheumatic, tumor, infectious diseases, drug-induced hemolysis.

If anemia appears several days or months after a blood transfusion or with prolonged jaundice in a newborn, a direct Coombs test is also performed.

Indirectantiglobulin test is performed before blood transfusion and during pregnancy of an Rh-negative woman.

Autoimmune hemolytic anemia

Autoimmune hemolytic anemia (primary)– a classic autoimmune disease with unknown reasons. The interaction within the immune system is disrupted, which leads to the perception of one’s own red blood cells as foreign. Antibodies are synthesized in lymph nodes IgG class(react at t 37°C) and/or IgM (at t 40°C), which, when attached to the surface of the erythrocyte, trigger a number of enzymes (the complement system) and “perforate” the wall of the erythrocyte, which leads to its destruction - hemolysis.

The first symptoms are caused by both the destruction of red blood cells and a decrease in hemoglobin. Among them:

• fatigue, general weakness, irritability
• dyspnea
• abdominal and chest pain, nausea
• dark urine color
• back pain
• icteric discoloration of the skin and mucous membranes
• decrease in the number of red blood cells and

Positive result direct Coombs tests 100% confirms the diagnosis of autoimmune hemolytic anemia, proving its autoimmune origin. Simultaneously negative result- does not make it possible to remove the diagnosis.

Secondary hemolytic anemia

Secondary autoimmune hemolytic anemia and a positive Coombs test can occur in the following diseases:

• Evans syndrome
• pneumonia infection

A positive antiglobulin test for these diseases is one of the symptoms, and not a criterion for diagnosis.

Hemolytic disease of the newborn

Cause hemolytic disease of newborns - incompatibility of the blood group of mother and fetus, in most cases according to the Rh system, in single cases - according to the AB0 system, casuistically - according to other antigens.

Rh conflict develops if the fetus of a Rh-negative woman inherits Rh-positive blood from the father.

The disease develops in a newborn only if the mother has already developed antibodies to the corresponding antigens, which happens after previous pregnancies, abortions, and incompatible blood transfusions. Most common reason triggering the synthesis of antibodies to antigens of the erythrocyte membrane - childbirth (feto-maternal bleeding). The first birth generally takes place without complications, but subsequent ones are fraught with hemolytic disease of the newborn in the first days after birth.

Symptoms of hemolytic disease of the newborn:

• yellowness of the skin
• , and mucous membranes
• enlarged liver and spleen
• breathing problems
• whole body swelling
• excitation and gradual depression of the central nervous system

Anemia after blood transfusion

Indirect Coombs test carried out before blood transfusion to assess compatibility, and a direct Coombs test - after it if post-transfusion hemolysis is suspected, i.e. in the presence of symptoms such as fever, watering (read below). The purpose of the analysis is to identify antibodies to transfused red blood cells that have bound to the red blood cells of the recipient and are the cause of post-transfusion hemolysis, as well as premature removal of donor red blood cells from the blood circulation of the recipient (the one who received the blood).

Symptoms:

• increase in body temperature
• skin rash
• back pain
• red
• nausea
• dizziness

Decoding

It is worth recalling that the fundamental rules for deciphering direct and indirect antiglobulin tests are the same. The only difference is the location of the antibodies - in the blood or on the red blood cell.

• If direct Coombs test is negative– this means that the antibody does not “sit” on the red blood cells and the cause of the symptoms should be sought further and an indirect Coombs test should be performed
• if a positive result of the Coombs test is detected after a blood transfusion, infections, drugs - positivity lasts up to 3 months (lifetime of red blood cells is 120 days - 3 months)
• a positive antiglobulin test result for an autoimmune disease lasts months or even years

Norm

• direct Coombs test - negative
• indirect Coombs test - negative

A qualitatively positive result is measured in the number of pluses from one to four (+, ++, +++, ++++), and quantitatively in digital form - 1:16, 1:256, etc.

Yes. Your doctor should definitely know that you have received a blood transfusion, since it affects the correct interpretation of test results now. When receiving someone else's (albeit tested many times) blood, there is always the possibility that your body will develop antibodies against the transfused blood. It is these antibodies that will have a negative impact on health. For subsequent blood transfusions, the doctor must know that you have already received transfusions, which means there has been time for the synthesis of antibodies. For pregnant women, this information is even more relevant.

3. If there is a mismatch in the Rh factor between mother and child, will all children be sick?

Depends on whether the child is Rh positive or negative (RhD). Carriers of blood groups I, II, III and IV can be either Rh positive or negative. In a situation where the mother is Rh negative and the child is Rh positive, antibodies will be produced already with the first pregnancy, but only after the first birth (or termination of pregnancy) will there be direct contact between the blood of the mother and the child. The hemolytic effect of antibodies will be realized only during the second and next birth, which will lead to hemolytic disease in the newborn.

Every woman with negative Rh factor should be carefully examined during pregnancy, and after childbirth preventive treatment to prevent the appearance of antibodies and further complications.

4. During pregnancy, is it necessary to know my husband’s blood type before performing a Coombs test?

You need to not only know, but also check the blood type of the child’s biological father during pregnancy.

Data

• first proposed in Cambridge in 1945
• sensitivity threshold - at least 300 fixed antibody molecules on one red blood cell
• the number of antibodies triggering hemolysis - individually for each person (from 16-30 to 300)
• dynamics of others laboratory parameters hemolytic anemia (hemoglobin, bilirubin, reticulocytes) may return to normal, and the Coombs test will remain at the same level

Coombs test was last modified: March 16th, 2018 by Maria Bodyan

Direct Coombs test. This test is used to prove the presence of blocking antibodies fixed in the child’s red blood cells. A positive direct test indicates sensitization and serves as a convincing sign of hemolytic disease of the newborn even before the appearance of other clinical signs. As an exception and only in very severe cases a direct Coombs test may be negative due to the already occurring, almost complete hemolysis of sensitized red blood cells.

A direct Coombs test is performed as follows: 5 drops of blood taken from the child’s heel are placed in a test tube and 5 ml is added saline solution. Stir well and centrifuge for 10 minutes. Separate clear liquid over the erythrocyte sediment. Then add 5 ml of saline again, mix and centrifuge. After mixing with saline three times, the red blood cells are well washed. After the last separation of the supernatant, the erythrocyte sediment in the amount of 0.1 ml is mixed with 0.9 ml of physiological solution. Apply 2-3 drops of this mixture to a glass slide and add one drop of Coombs serum. The presence of agglutination indicates that the reaction is positive (positive direct Coombs test). The study should be carried out at room temperature above 16° to avoid the effect of cold agglutinins.

Indirect Coombs test serves as evidence of the presence of free antibodies in maternal serum and is performed with maternal serum.

Hemolytic disease in a newborn with Rh incompatibility usually manifests itself after the second pregnancy. The first child is born healthy, the second with signs of mild anemia, and only after the third pregnancy are children born with clear signs hemolytic disease. Only pre-sensitized women can give birth to a child with symptoms of hemolytic disease during their first pregnancy. In some cases, immunization causes abortions and stillbirths. For the onset and severity of the disease, the condition of the placenta and the duration of exposure of the maternal agglutinins to the fetus are important. When agglutinins appear 10-14 weeks before birth, the child usually experiences subclinical forms. The early appearance of agglutinins, 15-26 weeks before birth, causes severe forms diseases. In all forms of the disease, the main process is hemolysis. The consequence of the antigen-antibody reaction is hemolysis, damage to the liver and brain capillaries. Depending on which lesion predominates, there are also various shapes diseases. Some anaphylactic phenomena are also dangerous. They lead to the formation of histamine-like substances, causing severe damage to the liver cells and especially to the ganglion cells of the basal ganglia, ammon's horn, medulla oblongata and even the cerebral cortex. When liver cells are damaged, hepatic jaundice is added to extrahepatic jaundice. Children die due to severe symptoms of kernicterus. If they survive, symptoms of damage to the nervous system remain (disorders of the extrapyramidal system with choreoathetotic movements, a peculiar dancing gait, forced movements of the head, sometimes a disorder of coordination of voluntary movements with frequent falls, increased tone muscles, mental retardation, i.e. with signs of the so-called. encephalopathia posticteria infantum).

on the use of the “Set of reagents for determining antigens and antibodies of the Rhesus system in human blood”
(AGS for Coombs test)

TU 9398-102-51203590-2012

RU No. RZN 2013/1255 dated 10/11/2013

I. INTRODUCTION

Standard serum for the Coombs test contains specific heteroimmune antibodies against human blood proteins and is intended for use in two reaction options - direct and indirect Coombs test.

Direct Coombs test used to determine the sensitization of erythrocytes in vivo in hemolytic disease of newborns and in patients with an autoimmune form of chronic hemolytic anemia, as well as in some other conditions. Carrying out a direct Coombs test consists in the fact that serum for the Coombs test is added to the red blood cells under study, previously washed from the proteins of their own plasma. If red blood cells have been sensitized in vivo, the reaction results in their agglutination.

Indirect Coombs test used as a test to identify the state of sensitization of the body, allowing the detection of antibodies that are in a free state in the blood serum of the person being tested; as a test for the compatibility of transfused blood, when the effect of the recipient's serum on the red blood cells of the intended donor is examined; and as a test for determining various group antigens in erythrocytes using preliminary exposure to standard sera containing antibodies of known specificity. Indirect Coombs test (all options) produced in two stages, the first of which is the incubation of the test (or standard) serum with standard (or test) erythrocytes, i.e. sensitization of erythrocytes in vitro. The second stage is the actual reaction with serum for the Coombs test, which is performed in the same way as the direct Coombs test.

The final result of the reaction in both direct and indirect Coombs tests occurs due to the interaction standard serum for the Coombs test, with antibodies (human blood protein) fixed on red blood cells. This result manifests itself as agglutination of red blood cells. To observe agglutination, you should use a white porcelain or any white plate with a wettable surface so that the drops applied to it are well mixed and do not spread over the surface of the plate. The mixed drops should be lightly spread over the plate, approximately to the size of a 2-kopeck coin.
II. PREPARATION OF REACTION INGREDIENTS

The test serum to determine the presence and specificity of antibodies in it is obtained from the blood of the person being examined without adding a preservative. Store at a temperature of + 4–8 0 C for no more than 2 days or frozen for 1 month.

Serum for compatibility testing is obtained in the same way, but is used with a shelf life of no more than 2 days.

Red blood cells intended for the determination of certain isoantigens in them are prepared with some kind of preservative from the person whose blood is being examined. You can also use red blood cell sediment from blood taken without a preservative.

When testing for compatibility, donor blood is used, drawn through a needle from a vial prepared for transfusion.

In all cases, red blood cells are washed from plasma 3-4 times in test tubes by adding 8-10 volumes of red blood cells to one volume isotonic solution sodium chloride, followed by stirring and centrifugation at 1500-2000 rpm for 5-10 minutes (until complete sedimentation of red blood cells). After each centrifugation, the supernatant is sucked off.
III. REACTION TECHNIQUES

The test erythrocytes are prepared in the form of a 5% suspension, for which one drop of erythrocytes washed four times is mixed in a test tube with 19 drops of isotonic sodium chloride solution.

One drop (0.05 ml) of a 5% suspension of red blood cells is transferred to a white plate and smeared. The plate is rocked and observed for 3 minutes to see if agglutination of red blood cells has occurred. If agglutination has not occurred, then add 1-2 drops of standard serum for the Coombs test, mix the drops thoroughly, then shake the plate slightly, then leave it alone for one or two minutes and shake again. At the same time, the result is monitored for 10 minutes, which is expressed in the presence or absence of agglutination.
Interpretation of the results of the direct Coombs test.

No agglutination – negative test

Presence of agglutination – a positive test, which indicates sensitization of the red blood cells being tested, i.e., the absorption of antibodies on them, which occurred in vivo in the human body (newborn, patient). If agglutination of the tested red blood cells occurred before adding serum for the Coombs test; the result of the direct test is not taken into account.
2. Indirect Coombs test

Intended for detection of isoantibodies in the blood serum of the examined person

It is used in cases where it is necessary to find out whether the blood of a patient, pregnant woman, donor, etc. contains isoimmune antibodies of incomplete form and to establish their specificity. For the reaction, the blood serum of the person being examined and standard red blood cells of known specificity are used. Among these 8-10 or more red blood cell samples there should be differences in factors from other systems and it is very important that each of these factors is contained in at least one red blood cell sample. The panel also includes samples of Rh-negative red blood cells with various combinations antigens of other systems, so that among them there are Duffy-positive and Duffy-negative, which, in turn, would be divided into Kell-positive and Kell-negative, Kidd-positive and Kidd-negative, etc. As a negative control, if possible, include erythrocytes of the person whose serum is being tested in the reaction.

Reaction technique

1 drop of a 5% suspension of erythrocytes from each test tube is transferred to a white plate, 1-2 drops of standard serum for the Coombs test are added there and the serum is thoroughly mixed with erythrocytes. The plate is slightly shaken, then left alone for 1-2 minutes and shaken periodically again, while simultaneously observing the result of the reaction for 20 minutes.

Interpretation of results

If in some or most drops, in addition to the control, observed agglutination , this means that the test serum contains incomplete antibodies against erythrocyte antigens. The question of the specificity of these antibodies is decided by comparing positive and negative reactions with the antigenic structure of red blood cells included in the reaction. The question of the activity of these antibodies is resolved by titration.

Example 1. Agglutination occurred with all samples of erythrocytes containing the Rh 0 (D) factor, regardless of the presence or absence of other factors of this system and other systems, while with all Rh 0 (D) - negative samples, agglutination was not observed (so regardless of the presence or absence of other factors) - this means that the test serum contains incomplete anti-Rhesus antibodies - Rh 0 (D).

Example 2. Agglutination occurred with all samples of erythrocytes containing the Duffy factor, regardless of the presence or absence of antigens of the Rhesus system and other systems, while no agglutination was observed with all Duffy-negative samples, which means that the test serum contains incomplete anti-antibodies. -Duffy.

Addition. If the institution does not have a complete panel of standard red blood cells, but there is a need to test serum for the presence of antibodies, it is recommended to conduct a study with 25-30 randomly taken samples of red blood cells healthy individuals group 0 (I) or the serum of the same name as the test serum. This will make it possible to resolve the issue of the presence or absence of antibodies to most antigens of the Rh-Hr, Duffy, Kell, Kidd system.

Upon receipt positive result further conclusion about the specificity of the detected antibodies can be decided upon special research with a complete panel of standard red blood cells.

3. Technique for performing an indirect Coombs test as a test for the compatibility of transfused blood

To prevent incompatibility during blood transfusion, the following compatibility tests must be performed before transfusion:

1. 
   A compatibility test for ABO blood groups, which is performed on a plane at room temperature.
2. 
   Compatibility tests for Rh factor and other isoantigens.

One small (0.01 ml) drop of washed donor erythrocytes, taken through a needle from a bottle of blood intended for transfusion, is transferred to the bottom of a centrifuge or other tube with a volume of 3-4 ml and 3 drops of the patient’s serum are added to it. The test tube is shaken to mix the erythrocytes with the serum, after which it is placed in a thermostat at 37 0 C for 45 minutes. After incubation, an isotonic sodium chloride solution is added to the top of the test tube, the contents of the test tube are mixed and centrifuged for 5-10 minutes. This washing of red blood cells is repeated 3 times, each time carefully removing the supernatant. When using centrifuge tubes, you can limit yourself to washing twice. Add 2 drops of isotonic sodium chloride solution to the washed red blood cells to obtain approximately 5% suspension.

1 drop of a 5% suspension of red blood cells is transferred to a white plate, 1-2 drops of standard serum for the Coombs test are added there and the serum is thoroughly mixed with red blood cells. Then the plate is slightly shaken, left alone for 1-2 minutes and periodically shaken again, while observing the result for 20 minutes.

Note: In case of Rh incompatibility, agglutination usually occurs within the first minute, but with a low titer of Rh antibodies (or other antibodies), agglutination may occur later, sometimes by the twentieth minute.
Interpretation of the result:

Agglutinates are visible in the form of lumps on a cleared or completely bleached background - this means that the donor’s blood incompatible with the recipient's blood and cannot be transfused to him.

Absence signs of agglutination means that the patient’s blood does not contain incomplete antibodies to the donor’s red blood cells in relation to the Rh factor and other isoantigens to which incomplete antibodies could be formed.

The indirect Coombs test is sensitive only for incomplete antibodies.
4. Indirect Coombs test technique for determining the antigenic structure of erythrocytes

This test is used to determine the presence or absence of any red blood cell group antigen using standard serum containing the corresponding antibodies in incomplete form.

Most often, the indirect Coombs test is used to determine the antigens Kell (K), Duffy (Fy), Kidd (Jk), and also to determine the weak antigen of the Rh system (Du).

Before proceeding with the determination of antigens, the red blood cells under study should be tested in a direct Coombs test. If the result of a direct test is positive, the determination of antigens by an indirect Coombs test cannot be carried out and other reactions must be used for this purpose.

For the reaction, centrifuge or other tubes with a volume of 3-4 ml are used in the amount of three: one for the test sample of erythrocytes and two for 1 small (0.01 ml) drop of three times washed test erythrocytes is added to the bottom of the first tube, control erythrocytes are added to the second tube, containing the desired antigen (for example, Duffy-positive) and into the third tube - negative control (Duffy-negative). Add 2-3 drops of standard serum (in this example, anti-Duffy) to all test tubes. The test tubes are shaken to mix the contents and placed in a thermostat at 37 0 C for 45 minutes. After such incubation, the test tubes are removed from the thermostat, an isotonic sodium chloride solution is added to the top, the contents are thoroughly mixed and centrifuged at 1500-2000 thousand rpm for 5-10 minutes. After centrifugation, the supernatant is carefully sucked off and this washing of red blood cells is repeated 4 times. (When using centrifuge tubes, you can limit yourself to washing twice).

Add 2 drops of isotonic sodium chloride solution to the washed red blood cells to obtain approximately 5% suspension. 1 drop of a 5% suspension of erythrocytes from each test tube is transferred to a white plate, 1-2 drops of standard serum for the Coombs test are added to each drop of erythrocytes and mixed. The plate is slightly shaken, then left alone for 1-2 minutes and shaken periodically again, while simultaneously observing the progress of the reaction for 20 minutes.
Interpretation of results

Presence of agglutination red blood cells (positive result) means that the blood contains the desired antigen (in this example, Duffy-positive blood).

No agglutination (negative result) means that the blood tested does not contain the desired antigen (Duffy negative).

The result is taken into account as true after checking the control samples, i.e. in the example, if the result is positive with a Duffy-positive sample and negative with a Duffy-negative sample.

5. Release form

The reagent is available in liquid form in bottles of 5 or 10 ml (1 ml contains 10 doses). Sodium azide is used as a preservative in a final concentration of 0.1%.

6.Storage

Shelf life: two years in the refrigerator at 2-8°C. The opened bottle is suitable for use when stored in the refrigerator in a sealed container. closed during the entire shelf life.

The grounds for complaint are: lack of activity, non-specificity, violation of the integrity of the bottle, presence of flakes, expired reagent. When making a complaint, please indicate the date of receipt, the supplier (if you received the product not from the manufacturer), the batch number, and the reasons why the reagent was found unsuitable. Please attach a protocol with the results of testing the reagent and 2-3 unopened bottles with the reagent
The complaint should be sent to the manufacturer LLC "MEDICLON": 127276

On the surface of red blood cells is a large number of antigens. Depending on the type of these antigens, blood groups are distinguished; the most studied groups are ABO, Rh, Kell, Duffy and many others...

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Description of the study

Preparing for the study: Blood is drawn from a vein and then serum (blood plasma without fibrinogen) is obtained by natural clotting or by fibrinogen precipitation. Test material: Taking blood

There are a large number of antigens on the surface of red blood cells. Depending on the type of these antigens, blood groups are distinguished; the most studied groups are ABO, Rh, Kell, Duffy and many other systems. Normally, there are antibodies to antigens of another group in the blood, but during blood transfusion, pregnancy, autoimmune diseases, etc. antibodies to their own antigens are detected. Incomplete antibodies to red blood cells

Method

The indirect Coombs reaction is based on the detection of agglutination (clumping) of red blood cells that have incomplete antibodies on the surface, which appears when antiglobulin serum is added.

At the first stage, donor red blood cells (O(I) group, Rh+) ​​and the test serum are added into one test tube. If incomplete antibodies to red blood cells are present in the test serum, then they are fixed on the surface of donor red blood cells.

At the second stage, donor red blood cells with antibodies (if any) and standard antiglobulin serum with antibodies to human immunoglobulins. If at the first stage, antibodies to red blood cells are fixed on the surface of red blood cells, then when standard serum is added, the red blood cells stick together due to the interaction of antibodies.

Reference values ​​- norm
(Indirect Coombs test (antiglobulin test, detection of incomplete anti-erythrocyte antibodies), blood)

Information regarding the reference values ​​of indicators, as well as the composition of the indicators included in the analysis, may differ slightly depending on the laboratory!

Norm:

Normally, there should be no antibodies to one’s own red blood cells; when the Coombs test is performed, red blood cell aggregation does not occur.

Indications

Research of humoral specific immunity if you suspect autoimmune reactions in the body, Rh conflict between mother and fetus, determination of blood compatibility between donor and recipient

Increasing values ​​(positive result)

Antibodies to red blood cells are detected when:

1. Autoimmune hemolytic anemia

2. Hemolytic disease newborns

3. Systemic diseases connective tissue

4. Chronic active hepatitis and etc.

The Coombs test is a specific laboratory test that detects antibodies located in or on the surface of a red blood cell. This procedure allows you to diagnose the immune system, including in newborns, as well as identify hemolytic transfusion reactions. The Coombs test is actively used in forensic medicine and scientific genetics to determine erythrocyte antigens. Compliance with all the rules for carrying out such an analysis allows you to obtain the most reliable result.

Purpose of the antiglobulin test

The direct Coombs test allows you to detect anti-erythrocyte antibodies that are fixed on red blood cells. A positive reaction in such a study indicates the development of an autoimmune disease. It should be noted that a negative result does not exclude the presence of antibodies, since antibodies are often found in free form, that is, they have no connection with red blood cells. In such cases, it is advisable to conduct an indirect Coombs test, which will allow the determination of autonomous substances in

How is the analysis carried out?

abor venous blood the patient is carried out in the morning on an empty stomach, despite the fact that no significant factors influencing the final result of such a test have been found. It is allowed to store the taken material at a temperature of 2 to 8 °C for no longer than seven days. In order for the indicators this study were as accurate as possible whole blood must be delivered to the laboratory within the first two hours. Ideally, the Coombs test should show a negative result, which indicates the absence of hemolytic changes in the body.

Decoding the final indicators

The Coombs test is a rather labor-intensive research method that requires careful and precise execution. When using such a test, there may be some difficulties that are associated with incorrect interpretation of the final results due to weak manifestations positive reactions. It should be noted that the unreliability of the analysis - namely, a positive Coombs test - may be a consequence of ineffective washing of red blood cells, contact with fatty
surface, as well as neutralization of antiglobulin reagents by components

serum. Another disadvantage of this research method is the instability of the taken material, the storage of which has certain features.

A false negative result may be caused by excessive shaking of the red blood cell suspension during resuspension. Erroneous results may also be due to the presence of contaminants of anti-complementary antibodies, which are adsorbed during incubation on the surface of the tested red blood cells, resulting in the appearance of a positive result. If the test samples are thoroughly washed and the reaction conditions are controlled, these shortcomings can be easily eliminated, which will increase the chances of obtaining the most reliable Coombs test values.