The meaning of blood groups according to the avo system. Genetics of blood groups and their polymorphisms. Errors due to technical reasons include
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Classification of human blood, depending on its characteristics, is of practical importance during surgical interventions that require its transfusion, in organ and tissue transplantology, in forensic medicine to establish the fact of paternity, maternity and in the event of the loss of children at an early age, and also for pregnancy planning.
A person’s group affiliation is determined by antigens located on the surface of red blood cells (erythrocytes), is an inherited trait and does not change throughout our lives. The world medical community recognizes different systems of human blood groups, but the generally accepted determination of blood group is the ABO system.
Classification
According to this system, blood is divided into subtypes O, A, B and AB, depending on the presence or absence of A and B antigens in it.
The discovery and study of group identification revealed an uneven distribution of A and B antigens among different races and nationalities of humanity. For example, the majority of residents of northern Europe are owners of antigen A. 80% American Indians have the first group, but the third and fourth do not occur in them. The indigenous people of Australia are people with the first group. And among the residents of Central and East Asia the third predominates.
This makes it possible for ethnographers to study the origin of existing races and peoples, to trace their settlement and migration around the planet.
How often do you get your blood tested?
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Only as prescribed by the attending physician 30%, 667 votes
Once a year and I think that’s enough 17%, 372 vote
At least twice a year 15%, 323 vote
More than twice a year but less than six times 11%, 248 votes
I take care of my health and rent once a month 7%, 151 voice
I'm afraid of this procedure and try not to pass 4%, 96 votes
21.10.2019
In addition, thanks to modern medical observations, a pattern has been established between the group identification of people and the frequency of certain diseases. These studies could lead to important medical discoveries.
Group 0
The first, or AB0, means that it does not contain antigens A or B. For a long time assumed that for this reason blood of this type could be transfused to all patients, regardless of their group affiliation, so its owners were called universal donors. According to research by anthropologists, it is the most ancient; its signs were found in primitive people who were engaged in hunting and gathering. 40-50% of the total population globe are representatives of this group subspecies.
It is believed that its carriers have a strong immune system, are less susceptible to infections, but more often than other people suffer from arthritis, allergies and peptic ulcer.
Group A
Red blood cells of the second blood group according to the AB0 system contain antigen A. They cannot be used as donor material for carriers of those groups where this antigen is absent.
It ranks second in prevalence - 30-40% of humanity. Strengths health - good metabolism and healthy digestion. Among antigen A carriers, disorders of the liver, gallbladder, cardiovascular diseases and diabetes are more often diagnosed.
Group B
In turn, red blood cells of the third blood group according to the ABO system contain B antigens, which are found only in 10-20% of the world population.
Important information: Does a person’s blood type and Rh factor change from positive to negative throughout life?
Among representatives of this class of humanity there is a tendency to develop chronic fatigue and the presence of autoimmune diseases, while agreeing that they are the owners of a strong and healthy digestive system.
Group AB
The blood of this species contains both A and B antigens, which is why its owners are called universal recipients.
It is the rarest, its carriers make up only 5% of the population. They have a strong immune system, but at the same time, various cardiovascular diseases.
Inheritance of group affiliation according to the ABO system occurs according to the classical laws of genetics:
- If the parents do not have antigens A, B, the child will not have them either.
- In families where parents (one or both) are holders of AB (IV) blood, a child with 0 blood cannot be born.
- If the mother and father have the second group, then the child will have the first or second.
Depending on the presence or absence of antigens A and B in a person’s red blood cells, his plasma may contain antibodies responsible for the destruction of foreign antigens. Any use of the recipient's blood or its components should be carried out only taking into account group compatibility with the donor.
In modern clinical practice transfusion of blood, red blood cells and plasma of the same type as the patient. In some emergency cases, group 0 red blood cells can be transfused into recipients of other subtypes. Group A red blood cells can be used for transfusion to patients of group A and AB, and red blood cells from donor B to B and AB recipients. We are talking only about red blood cells; the use of plasma and whole blood for patients of a different group can cause irreparable harm to their health.
| Donor blood | recipient | |||
| A | IN | AB | ||
| + | — | — | — | |
| A | — | + | — | — |
| IN | — | — | + | — |
| AB | — | — | — | + |
| Donor's red blood cells | recipient | |||
| A | IN | AB | ||
| + | + | + | + | |
| A | — | + | — | + |
| IN | — | — | + | + |
| AB | — | — | — | + |
To avoid complications during blood transfusion, even of the same group, a preliminary biological test is carried out: the patient is injected with 25 ml of donor material 3 times with breaks of 3 minutes, while monitoring the patient’s condition. Further transfusion of the required total amount of materials is carried out only if there are no signs of deterioration in the person’s condition.
How is a group defined?
To determine which ABO blood type a person is a carrier of, material taken from his finger is sufficient. Test reagents anti-A and anti-B are applied to a white plate, mixed with the test sample and the result is assessed after 3-5 minutes.
If clots form in the first sample, i.e. red blood cells stick together (agglutination), and in the second case, the red blood cells do not stick together, this means that the person has antigen A and does not have antigen B. In this case, the donor has the first group (A). Other groups are defined similarly.
AB0 blood groups
The blood groups of the AB0 system were discovered in 1900 by K. Landsteiner, who, by mixing the red blood cells of some individuals with the blood serum of other individuals, discovered that with some combinations the blood coagulates, forming flakes (agglutination reaction), but with others it does not. Based on these studies, Landsteiner divided the blood of all people into three groups: A, B and C. In 1907, another blood group was discovered.
It was found that the agglutination reaction occurs when antigens of one blood group (they are called agglutinogens), which are found in red blood cells - erythrocytes, stick together with antibodies of another group (they are called agglutinins) that are found in plasma - the liquid part of the blood. The division of blood according to the AB0 system into four groups is based on the fact that the blood may or may not contain antigens (agglutinogens) A and B, as well as antibodies (agglutinins) α (alpha or anti-A) and β (beta or anti-B) .
First blood group - 0 (I)
Group I - does not contain agglutinogens (antigens), but contains agglutinins (antibodies) α and β. It is designated 0 (I). Since this group does not contain foreign particles (antigens), it can be transfused to all people. A person with this blood type is a universal donor.
Second blood group A β (II)
Group II contains agglutinogen (antigen) A and agglutinin β (antibodies to agglutinogen B). Therefore, it can be transfused only to those groups that do not contain antigen B - these are groups I and II.
Third blood group Bα (III)
III groups a contains agglutinogen (antigen) B and agglutinin α (antibodies to agglutinogen A). Therefore, it can be transfused only to those groups that do not contain antigen A - these are groups I and III.
Fourth blood group AB0 (IV)
Blood group IV contains agglutinogens (antigens) A and B, but contains agglutinins (antibodies). Therefore, it can only be transfused to those who have the same, fourth blood group. But, since there are no antibodies in the blood of such people that can stick together with antibodies introduced from outside, they can be transfused with blood of any group. People with blood group IV are universal recipients.
Blood groups according to the ABO system
Method for determining blood groups
The blood group according to the ABO system is determined using an agglutination reaction. Currently, there are three ways to determine blood groups using the ABO system:
According to standard isohemagglutinating sera;
Using monoclonal antibodies (coliclones anti-A and anti-B).
Determination of blood groups using standard isohemagglutinating sera
The essence of the method is to detect group A and B antigens in the test blood using standard sera. For these purposes, the agglutination reaction is used. The test should be done in a room with good lighting at a temperature of 15-25 ° C.
To carry out the test you need: - Standard isohemagglutinating sera of groups O (I), A (II), B (III) and AB (IV) of two different series. Serums for determining blood groups are prepared from donor blood in special laboratories. Serums are stored in the refrigerator at a temperature of 4 - 8° C. The expiration date of the serum is indicated on the label. The label also indicates the titer (the maximum dilution of the serum at which an agglutination reaction can occur), which must be no lower than 1: 32 (for serum B (III) - no lower than 1:16 / 32). The whey should be transparent, without signs of rotting. For convenience, standard serums are tinted in a certain color: O (I) - colorless (gray), A (II) - blue, B (III) - red, AB (IV) - bright yellow. These colors accompany all labels on blood products that have a group affiliation (blood, red blood cells, plasma, etc.).
White porcelain or enamel plates or other plates with a wettable surface, marked according to blood types.
Isotonic sodium chloride solution.
Needles, pipettes, glass rods (slides).
Reaction technique.
1. Under the appropriate designations of the blood group, serum of groups I, II, III is applied to a plate (plate) in a volume of 0.1 ml (one large drop with a diameter of about 1 cm). To avoid mistakes, two series of sera are applied, since one of the series may have low activity and not give clear agglutination. Thus, 6 drops are obtained, forming two rows of three drops in the following order from left to right: 0 (I), A (II), B (III).
2. Blood for research is taken from a finger or from a vein. Using a dry glass rod, 6 drops of the test blood, approximately the size of a pinhead of 0.01 ml (small drop), are successively transferred to the plate at 6 points, each next to a drop of standard serum. Moreover, the amount of serum should be 10 times greater than the amount of blood being tested. Then they are carefully mixed together using glass rods with rounded edges.
More is possible simple technique: one large drop of blood is applied to the plate, then it is taken from there with the corner of a glass slide and each drop of serum is transferred, carefully mixing it with the last. In this case, the blood is taken each time with a clean corner of the glass, making sure that the drops do not mix.
3. After mixing the drops, shake the plate periodically. Agglutination begins within the first 10-30 seconds. But observation should be carried out for at least 5 minutes, since later agglutination is possible, for example with red blood cells of group A 2 (II).
4. Add one drop of isotonic sodium chloride solution to those drops where the reaction has occurred, after which the results of the reaction are assessed.
The agglutination reaction can be positive or negative
If the reaction is positive, within the first 10-30 seconds, small red grains (agglutinates), consisting of glued red blood cells, visible to the naked eye, are observed in the mixture. Small grains gradually merge into larger grains or even flakes irregular shape. In a negative reaction, the drop remains uniformly colored red.
The results of reactions of two series in drops with serum of the same group must coincide.
The belonging of the blood being tested to the corresponding group is determined by the presence or absence of agglutination when reacting with the corresponding sera.
If all serums were given positive reaction, which means that the tested blood contains both agglutinogens - A and B. But in such cases, in order to exclude a nonspecific agglutination reaction, additional control study test blood with standard serum of group AB (IV).
Determination of blood groups using monoclonal antibodies
To determine blood groups using this method, monoclonal antibodies are used, which are obtained using hybridoma biotechnology.
A hybridoma is a cell hybrid formed by the fusion of cells bone marrow(myeloma) with an immune lymphocyte that synthesizes specific monoclonal antibodies. The hybridoma has the ability to grow unlimitedly, is characteristic of a tumor cell, and has the inherent ability of a lymphocyte to synthesize antibodies.
Standard reagents have been developed: monoclonal antibodies (mAbs): anti-A and anti-B coliclones, used for the determination of erythrocyte agglutinogens. Zoliclones are lyophilized powder of red (anti-A) and blue (anti-B) color, which is diluted with isotonic sodium chloride solution immediately before the sample.
Technique.
Anti-A and anti-B zoliclones are applied to a white tablet, one large drop (0.1 ml) under the appropriate inscriptions: anti-A or anti-B. One small drop of the test blood should be applied next to the drops of antibodies. After mixing the components, observe the agglutination reaction for 2-3 minutes. Evaluating the results is very simple.
Determining blood grouping may be accompanied by errors that lead to incorrect interpretation of the results. Three main groups of errors can be distinguished: errors associated with low quality reagents; technical errors; errors associated with features Scheme for assessing the results of determining blood groups using monoclonal antibodies (coliclones anti-A and anti-B) of the blood being tested. The first two can be avoided by strictly observing the above-described requirements for serum, reaction conditions, etc. The third group is associated with the phenomenon of nonspecific panagglutination (Thomsen phenomenon), the essence of which is that serum at room temperature agglutinates with all red blood cells, even with their own (autoagglutination), and erythrocytes at the same time give agglutination with all sera, even with serum of group AB. A similar phenomenon has been described in a number of diseases: blood diseases, splenomegaly, liver cirrhosis, infectious diseases etc. Panagglutination and autoagglutination in healthy people have also been described.
The phenomenon of panagglutination and autoagglutination is observed only when room temperature. If the determination of group membership is carried out at a temperature of 37 ° C, they disappear.
It must be strictly remembered that in all cases of unclear or questionable results, blood groups should be re-determination using standard sera from other series, as well as in a cross-sectional manner.
66. Methods for determining blood groups using the Avo and Rh systems.
To conduct the study, standard hemagglutinating sera I (O), II (A), III (B), IV (AB) are required, and the first three serum options must be presented in two series.
The serum must be suitable for use; for this purpose, it is necessary to check its compliance with the expiration date indicated on the serum label, and visually determine its condition. You cannot use serum if it is cloudy, contains foreign impurities, flakes, suspension, or has changed color.
The serum is considered suitable for use if it is transparent, the ampoule has a label indicating its main properties (series, expiration date, group affiliation, color marking according to the group affiliation), the ampoule is not damaged or opened.
You need a clean plate, which must be divided into four parts, noting the correspondence to each specific blood group, a scarifier needle, sterile cotton swabs, a clean, dry, fat-free glass slide, and alcohol. Apply a drop of each serum to a plate in accordance with the labeling. Then the skin of the pad of the fourth finger of the left hand is treated with a sterile cotton swab with alcohol. Using a scarifier, the skin is pierced, removing the first drop of blood that appears (admixture of alcohol and tissue fluid to it can distort the results of the study). The next drop of blood is taken with the corner of a glass slide, for each drop of serum - with a clean corner of the glass. For research, add a drop of blood to a drop of hemagglutinating serum in a ratio of 10: 1. Then, carefully turning and shaking the plate, mix the blood. Agglutination is usually detected in the form of flakes that are clearly visualized. To clarify the result, an isotonic sodium chloride solution is added to the drop, after which the result is assessed with sufficient reliability.
One of the mandatory conditions for conducting the study is compliance with the temperature regime.
The optimal temperature is 20 – 25 °C, since already below 15 °C cold agglutination is observed, which sharply violates the specificity of this sample, and at a temperature environment above this interval, the rate of the agglutination reaction slows down sharply.
If hemagglutination occurs in a drop with sera I (O), III (B), but does not occur with serum II (A), and the result is similar with the sera of the two sera, this means that the blood being tested belongs to group III (B) according to the ABO system .
If hemagglutination occurs in a drop with serum I (O), II (A), but does not occur with serum III (B), this means that the blood being tested belongs to group II (A) according to the ABO system.
But a situation is also possible when hemagglutination does not occur with any of the test sera, and in both series. This means that the blood being tested does not contain agglutinogens and belongs to group I (O) according to the ABO system.
If agglutination occurs with all sera, from both series, this means that the test serum contains both agglutinogens (A and B) and belongs to group IV (AB) according to the ABO system.
Express method for determining blood group according to the Rh system. For the study, you must have a standard anti-Rhesus serum belonging to group IV (AB) according to the ABO system, a Petri dish, an isotonic sodium chloride solution, a 30% rheopolyglucin solution, and a clean, dry, fat-free glass slide. First, you need to dilute the standard anti-Rhesus serum with a solution of rheopolyglucin and apply a drop of it to a Petri dish. In addition, a drop of group IV (AB) serum, which does not contain antibodies, is applied to a Petri dish. Blood sampling is carried out similarly to the method described when determining the ABO system. A drop of blood is collected using the corner of a glass slide, added to a drop of serum and mixed gently. At another angle, take a drop to add to another serum and also mix. The preliminary result is assessed after 4 minutes, then a drop of isotonic sodium chloride solution must be added to each drop, and the final result is assessed after 2 minutes. If no agglutination flakes are observed in both drops, this indicates that the blood being tested is Rh negative. If agglutination occurred with both sera, the result of the study cannot be considered reliable. Finally, if agglutination occurred with the anti-Rh serum, but did not occur with the other, control serum, the blood is considered Rh-positive.
67. Modern rules of blood transfusion according to groups of the ABO system and the Rh system. Responsibilities of a blood transfusion physician
The need for infusion of blood or its components, as well as the choice of method and determination of the dosage of transfusion, are determined by the attending physician based on clinical symptoms and biochemical tests. The doctor performing the transfusion is obliged, regardless of the data from previous studies and tests, to personally carry out the following studies: determine the patient’s blood group according to the ABO system and compare the obtained data with the medical history; determine the donor’s blood type and compare the data obtained with the information on the container label; check the compatibility of the blood of the donor and the patient; obtain biological sample data. Transfusion of blood and its fractions that have not been tested for AIDS, serum hepatitis and syphilis is prohibited. Blood transfusion is carried out in compliance with all necessary aseptic measures. Infusion of blood or its fractions is allowed only if the Rh factor of the donor and recipient matches. If necessary, it is possible to inject Rh-negative blood of the first group into a person with any blood group in a volume of up to 0.5 liters (adults only). Rh-negative blood of the second and third groups can be transfused to a person with the second, third and fourth groups, regardless of the Rh factor. A person with blood group IV and positive Rh factor can be transfused with blood of any group. Erythrocyte mass of Rh-positive blood of the first group can be infused into a patient with any group with a Rh-positive factor. Blood of the second and third groups with an Rh-positive factor can be infused into a person with a fourth Rh-positive factor. One way or another, a compatibility test is required before transfusion. When immunoglobulins of rare specificity are detected in the blood, an individual approach to the selection of blood and specific compatibility tests are required.
Blood compatibility tests between donor and recipient before transfusion.
Finally, if it is determined that the donor’s blood and the recipient’s blood match the ABO and Rh systems, a biological compatibility test must be performed for transfusion. It is carried out from the very beginning of blood transfusion. Having connected the blood transfusion system, open the clamp and inject approximately 20 ml of donor blood in a stream, then close the clamp and carefully observe the patient's reaction for 3 minutes. Biological incompatibility between the blood of the donor and the patient is rare in clinical practice, but can be very dangerous to the health of the recipient. We can talk about its presence if facial hyperemia, psychomotor agitation are observed, the patient tries to get up, inappropriate behavior, tachycardia and tachypnea, and decreased blood pressure may be observed. Increased breathing may be combined with a feeling of shortness of breath. Such a reaction is absolute contraindication for transfusion of blood from a given donor to a given recipient. However, this does not exclude the use of other portions of donor blood from this person to other patients. If a biological test is not accompanied by the appearance of a similar reaction when repeated twice, it is considered suitable for transfusion. The transfusion continues, but throughout the entire time it is necessary to carefully monitor the recipient’s condition, his blood pressure, body temperature, general condition, frequency and rhythm of breathing, assess the frequency and quality of the pulse, pay attention to the patient’s subjective sensations: feeling of heat, feeling of lack of air, itching, pain in the lumbar region, chills, etc. Similar monitoring of the patient’s condition is carried out within 4 hours after blood transfusion , and if after this time there is no reaction to the transfusion, then the blood transfusion performed is considered successful, carried out without complications.
Determination of the AB0 blood group
Standard sera of blood group I II III IV of 2 series are used.
Wear an apron, mask, glasses, gloves, and sleeves.
Methodology.
1. Drop one drop of standard serums from 2 series (6 drops) onto a clean, dry, fat-free plate. Each serum is dropped into a separate pipette. The pipettes are labeled and are placed in bottles with saline solution.
2. Next to a drop of standard serum, a drop of blood is applied with a stick or a separate pipette (serum-to-blood ratio 10:1).
3. The drops are thoroughly mixed with separate sticks.
4. Rock the plate and observe the results for 5 minutes. After 3 minutes, to exclude false agglutination, add a drop of saline solution and continue observation.
Definition of results.
Sera of all groups did not give a reaction; the blood tested belongs to group 0(I)
Sera of groups 0 (I) and B (III) gave a positive reaction, serum of groups A (II) did not cause agglutination of erythrocytes - the blood tested belongs to group A (II)
Sera of groups O (I) and A (II) gave a positive reaction, serum of group B (III) did not give a reaction - the blood is assigned to the group
Sera of all groups gave a positive reaction - the blood can be classified as group AB (IV). but only if, when testing the red blood cells under study with standard serum of group AB (IV), there is no agglutination reaction. Positive agglutination with AB (IV) serum means the presence of nonspecific agglutination.
Determination of ABO blood group using monoclonal antibodies.
This modern method determination of the ABO blood group using monoclonal reagents (anti-A and anti-B coliclones).
Determination method: Put on an apron, mask, gloves, arm ruffles
1. drop 1 drop of anti-A and anti-B cyclones onto a plate in two cells
2. Next to the drops of antibodies, the test blood is applied in a serum-to-blood ratio of 10:1.
3. Mix the reagents using separate glass rods.
4. Observe the result for 2.5 minutes.
Definition of results:
There is no agglutination with either tsoliclone anti-A or tsoliclone anti-B-
blood belongs to group O(I)
Agglutination is observed only with anti-A coliclone - blood belongs to group A (II)
Agglutination is observed only with anti-B coliclone - blood belongs to group B (III)
Agglutination is observed with both tsoliclone anti-A and tsoliclone anti-B - the blood belongs to group AB (IV)
5. Samples before blood transfusion.
Wear a mask, glasses, apron, gloves.
5 ml of blood is taken from the patient’s vein, centrifuged and serum is obtained.
A) Test for individual compatibility: apply 2-3 drops of serum to the tablet and add a drop of donor blood (10:1). The blood is mixed with the serum with a dry glass rod, the tablet is slightly rocked and the result is observed.
Result: absence of agglutination - the blood is compatible; there is agglutination - the blood is not compatible.
B) Test for Rh compatibility:
Write on a clean test tube the last name, first name, patronymic of the recipient and his blood type, as well as the number of the donor vial.
Add 2 drops of recipient serum, 1 drop of donor blood and 1 drop of 33% polyglucin solution to the bottom of the test tube using a Pasteur pipette.
Mix the mixture obtained in the test tube and tilt the test tube until horizontal position, turn it so that the contents spread along the walls of the test tube in its lower third.
After 5 minutes, add 2-3 ml of isotonic sodium chloride solution into the test tube and mix the contents by inverting the test tube 2-3 times.
Result:
1. The presence of red blood cell agglutination - the donor’s blood is incompatible with the patient’s blood.
2. There are no signs of agglutination - the blood is compatible.
B) Biological sample.
Technique:
Streamly transfuse 10-15 ml of blood (erythrocyte mass, plasma), then observe the patient for 3 minutes. With absence clinical manifestations reactions or complications (increased heart rate, breathing, shortness of breath, difficulty breathing and facial flushing, etc.), reintroduce 10-15 ml of blood (erythrocyte mass, plasma, etc.) and observe the patient for 3 minutes . The same procedure is repeated a third time. Lack of response in the patient after 3-fold testing is grounds for continuing the transfusion. 10-15 ml of blood is left in a vial and stored for 1 day in the refrigerator.
6.After blood transfusion The recipient is kept in bed for 2 hours and is observed by a doctor. His body temperature and blood pressure are measured hourly, and these readings are recorded in the medical history. The presence of urination and normal urine color are monitored. The next day after the transfusion, it is necessary to perform general analysis urine and blood. The indication for blood transfusion is recorded in the medical history and a transfusion protocol is drawn up containing the following information:
– Full name of the patient
Blood type and Rh status of the patient
Passport data of blood bottles or components, surname and initials of the donor, blood group and Rh, bottle number, date of procurement.
The result of the control determination of the patient’s blood group
Results of control determination of the group of each bottle of blood and its components.
Results of blood compatibility tests between donors and recipient at room temperature
Method and results of temperature compatibility tests
Biological sample results
The patient's condition during and after the transfusion.
/ 29
29 Methods for determining blood groups and Rh factor. Prevention of possible errors
Determination of blood groups using standard sera. One large drop of standard serum of groups 0(1), A(P), V(III) of two different series of each group is sequentially pipetted into the wells of the plate for determining blood groups. Then a drop of the test blood, 5 to 10 times less than the serum, is applied to each eyepiece. Using dry glass rods or by shaking the tablet, mix standard serum and test blood. After 5 minutes, the result is assessed (Table 2).
table 2
Determination of blood groups using standard sera
|
Test blood group |
Standard serums |
||
Determination of blood groups using standard red blood cells. Large drops of plasma from the blood being tested are placed into the wells of the plate. Next to each drop of plasma, a small drop of standard red blood cells of groups 0(1), A(II), B(III) is sequentially applied. The drops are mixed by shaking the tablet. After 5 minutes, add 1 drop of saline solution to each drop of the mixture and take into account the result.
Table 3
Determination of blood groups using standard red blood cells
|
Test blood group |
Standard red blood cells |
||
Determination of the Rh factor. Apply 1 large drop of the universal anti-Rhesus reagent to the bottom of a dry test tube, then add the same drop of the blood being tested. After shaking the tube, the anti-Rh serum and blood are mixed along the walls of the tube for 5 minutes, after which 5-7 ml of saline is added. The presence of agglutination indicates the presence of the Rh factor.
Determination of blood groups using cyclones. Coliclones anti-A, anti-B and anti-AB are intended for determining human blood groups of the ABO system in direct hemagglutination reactions and are used instead of or in parallel with polyclonal immune sera.
Monoclonal anti-A and anti-B antibodies are produced by two mouse hybridomas and belong to class M immunoglobulins. Coliclones are prepared from the ascites fluid of mice carrying anti-A and anti-B hybridomas. Coliclone anti-AB is a mixture of monoclonal anti-A and anti-B antibodies.
Tsoliklons are available in liquid form in bottles of 5 - 10 ml. Tsoliclone anti-A - yellow color, anti-B - blue, anti-AB - colorless. Sodium azide is used as a preservative.
Technique for determining blood groups. The determination is made in native blood taken as a preservative; in blood taken without a preservative, including from a finger. The method of direct hemagglutination on a plate or tablet is used, the determination is carried out in a room with good lighting at a temperature of 15 - 25 X.
Anti-A, anti-B and anti-AB zoliclones are applied to a plate or tablet with individual pipettes, one large drop (0.1 ml) under the appropriate inscriptions.
One small drop of test blood (0.01 - 0.03 ml) is applied next to the drops of antibodies.
Mix the blood with the reagent by shaking the tablet.
Observe the progress of the reaction with zoliclones visually by gently shaking the plate for 3 minutes. Agglutination of erythrocytes with coliclones usually occurs in the first 3 to 5 seconds, but observation should be carried out for 3 minutes due to the later appearance of agglutination with erythrocytes containing weak varieties of antigens A or B.
The result of the reaction in each drop can be positive or negative. A positive result is expressed in agglutination (sticking together) of red blood cells. Aglutinates are visible as small red aggregates that quickly mix into large flakes. In a negative reaction, the drop remains uniformly colored red, and no agglutinates are detected in it.
Interpretation of the results of the agglutination reaction with zoliclones is presented in table. 4.
Table 4
Determination of blood groups using zoliclones
|
Test blood group |
Standard red blood cells |
||
Note: the sign (+) indicates the presence of agglutination, the sign (-) indicates its absence.
If the result of the agglutination reaction with all rattling zoliclones is positive, it is necessary to exclude spontaneous nonspecific agglutination of the tested erythrocytes. To do this, mix 1 drop of the red blood cells under study with a drop of physiological solution on a plane. Blood can be classified as group AB(IV) only if there is no agglutination of red blood cells in saline solution.
Determination of the Rh factor using zoliclon. Apply 1 large drop of anti-D-super zoliclone into the lens of the tablet and apply 1 small drop of the test blood next to it.
Shaking the tablet, mix the blood with the zoliclon. The progress of the reaction is monitored for 3 minutes. The presence of agglutination indicates the presence of the Rh factor.
Functions. Blood groups are genetically inherited characteristics that do not change throughout life. natural conditions. A blood group is a specific combination of surface antigens of erythrocytes (agglutinogens) of the ABO system. Determination of group membership is widely used in clinical practice during transfusion of blood and its components, in gynecology and obstetrics when planning and managing pregnancy. The AB0 blood group system is the main system that determines the compatibility and incompatibility of transfused blood, because its constituent antigens are the most immunogenic. A feature of the AB0 system is that in the plasma of non-immune people there are natural antibodies to an antigen that is absent on red blood cells. The AB0 blood group system consists of two group erythrocyte agglutinogens (A and B) and two corresponding antibodies - plasma agglutinins alpha (anti-A) and beta (anti-B). Various combinations of antigens and antibodies form 4 blood groups:
- Group 0(I) - there are no group agglutinogens on red blood cells, agglutinins alpha and beta are present in the plasma.
- Group A (II) - red blood cells contain only agglutinogen A, agglutinin beta is present in the plasma;
- Group B (III) - red blood cells contain only agglutinogen B, plasma contains agglutinin alpha;
- Group AB (IV) - antigens A and B are present on red blood cells, plasma does not contain agglutinins.
Determination of blood groups is carried out by identifying specific antigens and antibodies (double method, or cross reaction).
Blood incompatibility is observed if the red blood cells of one blood carry agglutinogens (A or B), and the plasma of another blood contains the corresponding agglutinins (alpha or beta), and an agglutination reaction occurs.
Transfusion of red blood cells, plasma and especially whole blood from a donor to a recipient must be strictly observed group compatibility. To avoid blood compatibility donor and recipient, it is necessary laboratory methods accurately determine their blood types. It is best to transfuse blood, red blood cells and plasma of the same group as determined for the recipient. In emergency cases, group 0 red blood cells (but not whole blood!) can be transfused into recipients with other blood groups; Group A red blood cells can be transfused into recipients with blood group A and AB, and red blood cells from a group B donor can be transfused into group B and AB recipients.
Blood group compatibility cards (agglutination is indicated by a + sign):
Donor blood | Recipient's blood |
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Donor's red blood cells | Recipient's blood |
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Group agglutinogens are found in the stroma and membrane of erythrocytes. Antigens of the ABO system are detected not only on red blood cells, but also on cells of other tissues or can even be dissolved in saliva and other body fluids. They develop on early stages intrauterine development, and in the newborn are already present in significant quantities. The blood of newborn children has age-related characteristics - characteristic group agglutinins may not yet be present in the plasma, which begin to be produced later (constantly detected after 10 months) and the determination of the blood group in newborns in this case is carried out only by the presence of antigens of the ABO system.
In addition to situations involving the need for blood transfusion, determination of blood type, Rh factor, as well as the presence of alloimmune anti-erythrocyte antibodies should be carried out when planning or during pregnancy to identify the likelihood of an immunological conflict between mother and child, which can lead to hemolytic disease newborns.
Hemolytic disease of the newborn
Hemolytic jaundice of newborns, caused by an immunological conflict between mother and fetus due to incompatibility of erythrocyte antigens. The disease is caused by incompatibility of the fetus and mother for D-Rhesus or ABO antigens, less often there is incompatibility for other Rhesus (C, E, c, d, e) or M-, M-, Kell-, Duffy-, Kidd- antigens. Any of these antigens (usually D-Rh antigen), penetrating into the blood of a Rh-negative mother, causes the formation of specific antibodies in her body. The latter enter the fetal blood through the placenta, where they destroy the corresponding antigen-containing red blood cells. Predispose to the development of hemolytic disease of the newborn by impaired placental permeability, repeated pregnancies and blood transfusions to a woman without taking into account the Rh factor, etc. With early manifestations of the disease, an immunological conflict can cause premature birth or miscarriages.
There are varieties (weak variants) of antigen A (to a greater extent) and less frequently of antigen B. As for antigen A, there are options: “strong” A1 (more than 80%), weak A2 (less than 20%), and even weaker ones (A3 , A4, Ah - rarely). This theoretical concept is important for blood transfusion and can cause accidents when assigning donor A2 (II) to group 0 (I) or donor A2B (IV) to group B (III), since the weak form of antigen A sometimes causes errors in the determination blood groups of the ABO system. Correct identification of weak A antigen variants may require repeated testing with specific reagents.
Decrease or complete absence natural agglutinins alpha and beta are sometimes noted in immunodeficiency states:
- neoplasms and blood diseases - Hodgkin's disease, multiple myeloma, chronic lymphatic leukemia;
- congenital hypo- and agammaglobulinemia;
- in children early age and in the elderly;
- immunosuppressive therapy;
- severe infections.
Difficulties in determining the blood group due to suppression of the hemagglutination reaction also arise after the introduction of plasma substitutes, blood transfusion, transplantation, septicemia, etc.
Inheritance of blood groups
The laws of inheritance of blood groups are based on the following concepts. There are three possible variants (alleles) at the ABO gene locus - 0, A and B, which are expressed in an autosomal codominant manner. This means that individuals who have inherited genes A and B express the products of both of these genes, resulting in the AB (IV) phenotype. Phenotype A (II) can be present in a person who has inherited from parents either two genes A, or genes A and 0. Accordingly, phenotype B (III) - when inheriting either two genes B, or B and 0. Phenotype 0 (I) appears when inheritance of two genes 0. Thus, if both parents have blood group II (genotypes AA or A0), one of their children may have the first group (genotype 00). If one of the parents has blood type A(II) with a possible genotype AA and A0, and the other has B(III) with a possible genotype BB or B0, children can have blood groups 0(I), A(II), B(III) ) or AB (!V).
- Hemolytic disease of newborns (detection of incompatibility between the blood of mother and fetus according to the AB0 system);
- Preoperative preparation;
- Pregnancy (preparation and follow-up of pregnant women with negative Rh factor)
Preparation for the study: not required
If necessary (detection of the A2 subtype), additional testing is carried out using specific reagents.
Execution time: 1 day
Research result:
- 0 (I) - first group,
- A (II) - second group,
- B (III) - third group,
- AB (IV) - fourth blood group.
Rh factor Rh
The main surface erythrocyte antigen of the Rh system, by which a person’s Rh status is assessed.
Functions. Rh antigen is one of the erythrocyte antigens of the Rh system, located on the surface of erythrocytes. There are 5 main antigens in the Rh system. The main (most immunogenic) antigen is Rh (D), which is usually referred to as the Rh factor. The red blood cells of approximately 85% of people carry this protein, so they are classified as Rh positive (positive). 15% of people do not have it and are Rh negative (Rh negative). The presence of the Rh factor does not depend on group membership according to the AB0 system, does not change throughout life, does not depend on external reasons. It appears in the early stages of intrauterine development, and is already found in a significant amount in the newborn. Determination of Rh blood is used in general clinical practice during transfusion of blood and its components, as well as in gynecology and obstetrics when planning and managing pregnancy.
Incompatibility of blood according to the Rh factor (Rh conflict) during blood transfusion is observed if the donor's red blood cells carry Rh agglutinogen, and the recipient is Rh negative. In this case, the Rh-negative recipient begins to produce antibodies directed against the Rh antigen, leading to the destruction of red blood cells. Transfusions of red blood cells, plasma, and especially whole blood from a donor to a recipient must strictly observe compatibility not only by blood type, but also by Rh factor. The presence and titer of antibodies to the Rh factor and other alloimmune antibodies already present in the blood can be determined by specifying the “anti-Rh (titer)” test.
Determination of blood type, Rh factor, and the presence of alloimmune anti-erythrocyte antibodies should be carried out when planning or during pregnancy to identify the likelihood of an immunological conflict between mother and child, which can lead to hemolytic disease of the newborn. The occurrence of Rh conflict and the development of hemolytic disease of newborns is possible if the pregnant woman is Rh negative and the fetus is Rh positive. If the mother is Rh + and the fetus is Rh negative, there is no danger of hemolytic disease for the fetus.
Hemolytic disease of the fetus and newborns- hemolytic jaundice of newborns, caused by an immunological conflict between mother and fetus due to incompatibility of erythrocyte antigens. The disease can be caused by incompatibility of the fetus and mother for D-Rhesus or ABO antigens, less often there is incompatibility for other Rhesus (C, E, c, d, e) or M-, N-, Kell-, Duffy-, Kidd antigens (according to statistics, 98% of cases of hemolytic disease of newborns are associated with D - Rh antigen). Any of these antigens, penetrating the blood of a Rh-negative mother, causes the formation of specific antibodies in her body. The latter enter the fetal blood through the placenta, where they destroy the corresponding antigen-containing red blood cells. Predisposition to the development of hemolytic disease of newborns is impaired placental permeability, repeated pregnancies and blood transfusions to a woman without taking into account the Rh factor, etc. With early manifestations of the disease, an immunological conflict can cause premature birth or repeated miscarriages.
Currently, there is a possibility of medical prevention of the development of Rh conflict and hemolytic disease of newborns. All Rh-negative women during pregnancy should be under medical supervision. It is also necessary to monitor the level of Rh antibodies over time.
There is a small category of Rh-positive individuals who are able to form anti-Rh antibodies. These are individuals whose red blood cells are characterized by significantly reduced expression of the normal Rh antigen on the membrane (“weak” D, Dweak) or expression of an altered Rh antigen (partial D, Dpartial). In laboratory practice, these weak variants of the D antigen are combined into the Du group, the frequency of which is about 1%.
Recipients containing Du antigen should be classified as Rh-negative and should be transfused only with Rh-negative blood, since normal D antigen can cause an immune response in such individuals. Donors with the Du antigen qualify as Rh-positive donors, since transfusion of their blood can cause an immune response in Rh-negative recipients, and in the case of previous sensitization to the D antigen, severe transfusion reactions.
Inheritance of the Rh blood factor.
The laws of inheritance are based on the following concepts. The gene encoding the Rh factor D (Rh) is dominant, the allelic gene d is recessive (Rh-positive people can have the DD or Dd genotype, Rh-negative people can only have the dd genotype). A person receives 1 gene from each parent - D or d, and thus has 3 genotype options - DD, Dd or dd. In the first two cases (DD and Dd), a blood test for Rh factor will give a positive result. Only with the dd genotype will a person have Rh negative blood.
Let's consider some variants of the combination of genes that determine the presence of the Rh factor in parents and children
- 1) The father is Rh positive (homozygote, genotype DD), the mother is Rh negative (genotype dd). In this case, all children will be Rh positive (100% probability).
- 2) The father is Rh positive (heterozygote, genotype Dd), the mother is Rh negative (genotype dd). In this case, the probability of having a child with negative or positive Rh is the same and equal to 50%.
- 3) The father and mother are heterozygotes for this gene (Dd), both are Rh positive. In this case, it is possible (with a probability of about 25%) to give birth to a child with negative Rh.
Indications for the purpose of analysis:
- Determination of transfusion compatibility;
- Hemolytic disease of newborns (detection of incompatibility between the blood of mother and fetus according to the Rh factor);
- Preoperative preparation;
- Pregnancy (prevention of Rh conflict).
Preparation for the study: not required.
Material for research: whole blood (with EDTA)
Determination method: Filtration of blood samples through a gel impregnated with monoclonal reagents - agglutination + gel filtration (cards, crossover method).
Execution time: 1 day
Interpretation of results:
The result is given in the form:
Rh + positive Rh - negative
When weak subtypes of antigen D (Du) are detected, a comment is issued: “a weak Rh antigen (Du) has been detected, it is recommended to transfuse Rh-negative blood if necessary.”
Anti-Rh (alloimmune antibodies to the Rh factor and other erythrocyte antigens)
Antibodies to the clinically most important erythrocyte antigens, primarily the Rh factor, indicating the body's sensitization to these antigens.
Functions. Rh antibodies belong to the so-called alloimmune antibodies. Alloimmune anti-erythrocyte antibodies (to the Rh factor or other erythrocyte antigens) appear in the blood under special conditions - after a transfusion of immunologically incompatible donor blood or during pregnancy, when fetal red blood cells carrying paternal antigens that are immunologically foreign to the mother penetrate through the placenta into the woman’s blood. Non-immune Rh-negative people do not have antibodies to the Rh factor. In the Rh system, there are 5 main antigens, the main (most immunogenic) is antigen D (Rh), which is usually referred to as the Rh factor. In addition to the Rh system antigens, there are a number of clinically important erythrocyte antigens to which sensitization can occur, causing complications during blood transfusion. The method of screening blood for the presence of alloimmune anti-erythrocyte antibodies, used in INVITRO, allows, in addition to antibodies to the Rh factor RH1(D), to detect alloimmune antibodies to other erythrocyte antigens in the test serum.
The gene encoding the Rh factor D (Rh) is dominant, the allelic gene d is recessive (Rh-positive people can have the DD or Dd genotype, Rh-negative people can only have the dd genotype). During pregnancy of a Rh-negative woman with a Rh-positive fetus, the development of an immunological conflict between mother and fetus due to the Rh factor is possible. Rh conflict can lead to miscarriage or the development of hemolytic disease of the fetus and newborns. Therefore, determination of blood type, Rh factor, as well as the presence of alloimmune anti-erythrocyte antibodies should be carried out when planning or during pregnancy to identify the likelihood of an immunological conflict between mother and child. The occurrence of Rh conflict and the development of hemolytic disease of newborns is possible if the pregnant woman is Rh negative and the fetus is Rh positive. If the mother has a positive Rh antigen and the fetus is negative, a conflict regarding the Rh factor does not develop. The incidence of Rh incompatibility is 1 case per 200-250 births.
Hemolytic disease of the fetus and newborns is hemolytic jaundice of newborns, caused by an immunological conflict between mother and fetus due to incompatibility of erythrocyte antigens. The disease is caused by incompatibility of the fetus and mother for D-Rhesus or ABO (group) antigens, less often there is incompatibility for other Rhesus (C, E, c, d, e) or M-, M-, Kell-, Duffy- , Kidd antigens. Any of these antigens (usually D-Rh antigen), penetrating into the blood of a Rh-negative mother, causes the formation of specific antibodies in her body. The penetration of antigens into the maternal bloodstream is facilitated by infectious factors that increase the permeability of the placenta, minor injuries, hemorrhages and other damage to the placenta. The latter enter the fetal blood through the placenta, where they destroy the corresponding antigen-containing red blood cells. Predisposition to the development of hemolytic disease of newborns is impaired placental permeability, repeated pregnancies and blood transfusions to a woman without taking into account the Rh factor, etc. With early manifestations of the disease, an immunological conflict can cause premature birth or miscarriages.
During the first pregnancy with a Rh-positive fetus, a pregnant woman with Rh "-" has a 10-15% risk of developing a Rh conflict. The first meeting of the mother's body with a foreign antigen occurs, the accumulation of antibodies occurs gradually, starting from approximately 7-8 weeks of pregnancy. The risk of incompatibility increases with each subsequent pregnancy with a Rh-positive fetus, regardless of how it ended (artificial abortion, miscarriage or childbirth, surgery for an ectopic pregnancy), bleeding during the first pregnancy, manual separation placenta, as well as if childbirth is carried out by cesarean section or is accompanied by significant blood loss. with transfusions of Rh-positive blood (if they were carried out even in childhood). If a subsequent pregnancy develops with an Rh-negative fetus, incompatibility does not develop.
All pregnant women with Rh "-" are placed on special registration in the antenatal clinic and dynamic monitoring of the level of Rh antibodies is carried out. For the first time, an antibody test must be taken from the 8th to the 20th week of pregnancy, and then periodically check the antibody titer: once a month until the 30th week of pregnancy, twice a month until the 36th week and once a week until the 36th week. Termination of pregnancy at less than 6-7 weeks may not lead to the formation of Rh antibodies in the mother. In this case, during a subsequent pregnancy, if the fetus has a positive Rh factor, the probability of developing immunological incompatibility will again be 10-15%.
Testing for alloimmune anti-erythrocyte antibodies is also important in general preoperative preparation, especially for people who have previously received blood transfusions.
Indications for the purpose of analysis:
- Pregnancy (prevention of Rh conflict);
- Monitoring of pregnant women with negative Rh factor;
- Miscarriage;
- Hemolytic disease of newborns;
- Preparation for blood transfusion.
Preparation for the study: not required.
Material for research: whole blood (with EDTA)
Determination method: agglutination + gel filtration method (cards). Incubation of standard typed erythrocytes with the test serum and filtration by centrifugation of the mixture through a gel impregnated with a polyspecific antiglobilin reagent. Agglutinated red blood cells are detected on the surface of the gel or in its thickness.
The method uses suspensions of erythrocytes from group 0(1) donors, typed according to erythrocyte antigens RH1(D), RH2(C), RH8(Cw), RH3(E), RH4(c), RH5(e), KEL1( K), KEL2(k), FY1(Fy a) FY2(Fy b), JK (Jk a), JK2(Jk b), LU1 (Lu a), LU2 (LU b), LE1 (LE a), LE2 (LE b), MNS1(M), MNS2 (N), MNS3 (S), MNS4(s), P1 (P).
Execution time: 1 day
When alloimmune anti-erythrocyte antibodies are detected, their semi-quantitative determination is carried out.
The result is given in titers (the maximum dilution of the serum at which a positive result is still detected).
Units of measurement and conversion factors: U/ml
Reference values: negative.
Positive result: Sensitization to Rh antigen or other erythrocyte antigens.
Blood groups.
Blood groups- normal immunogenetic blood characteristics that allow people to be grouped into certain groups according to
similarity of blood antigens.
Knowledge about blood groups underlies the doctrine of transfusion, organ and tissue transplantation, and forensic medicine.
Antigens - substances that enter the body parenterally and cause a specific immunological reaction, which manifests itself in the production of specific antibodies.
Antibodies - proteins of the globulin fraction of blood serum, which are formed in response to the introduction of an antigen and specifically interact with the antigens that caused their formation.
There are more than 250 isoantigens found in genetic connection. The following are important in the clinic:
1. AVO system
3. Lutheran
4. Kell-Cellano
AVO system.
The systematic study of blood groups began with the discovery of the blood groups of the ABO system by Landsteiner in 1901.
In this system, human red blood cells are divided according to the principle of having three different antigenic properties: A, B and AB (A+B). The antigenic properties of "O" do not exist; in extreme cases, we can talk about property H, but specific antibodies anti-N have very little clinical significance. Thus, a person’s blood group is determined by the antigenic properties of red blood cells.
As a rule, there are no ABO antibodies in the blood of a newborn. During the first year of life, the child develops antibodies to those antigens that are not in his own red blood cells (isoantigens, anti-A, anti-B). After this, serum, for example, group O, contains anti-A and anti-B antibodies, and serum of group AB does not contain these antibodies.
It has now been established that the intestines contain bacteria that carry the same antigenic determinants as red blood cells: the so-called heterophilic antigens. Most antibodies of the ABO system are of the type
class M immunoglobulins. Possessing 10 antigen binding sites, they are complete antibodies capable of causing agglutination of red blood cells.
Inheritance of blood groups.
The diploid set of chromosomes of each person contains 2 of the three allelic genes - A, B and 0 (H), encoding the properties of blood elements.
It has been shown that properties A and B are dominant, therefore blood group 0 is phenotypically expressed only in homozygotes. Since the AO and BO genotypes can produce phenotypes A and B, respectively, parents with one of these blood groups may well have a child with blood group 0. Alleles A and B are in a codominance relationship: in the presence of both of these genes, each of them is expressed, not interacting with each other.
Knowing these principles of inheritance, you can get some information about the parents based on the child's blood type. In forensic medical practice, it is generally accepted that a man with group AB cannot be the father of a child with blood group 0. The more group factors are taken into account, the more reliably paternity can be excluded (at present, a probability of 99% can be reached).
Blood typeA divided into subgroups A1 and A2. The main difference between these subgroups is that when mixed with anti-A serum, A1 red blood cells agglutinate faster and to a greater extent than A2. There are more H-structures on group A2 red blood cells than on A1 red blood cells. Approximately 80% of people with blood type A belong to the A1 subgroup, the remaining 20% belong to the A2 subgroup. This division has no practical significance for blood transfusion, since transfusion reactions between subgroups A1 and A2 are weakly expressed and are rare. Geographic distribution of blood groups.
More than 40% of residents Central Europe have blood group A, approximately 40% have group 0; 10% or more are group B and about 6% are group AB. 90% of Native Americans have group 0. More than 20% of the population Central Asia have blood type B. Based on data on the presence and correlation of various blood groups in certain areas of the globe, anthropologists can draw conclusions about the origin and mixing of peoples.
Rhesus (Rh) system.
Most Europeans are Rh positive (Rh+). This means that if their blood is mixed with the serum of rabbits previously immunized with rhesus monkey red blood cells, agglutination will occur. If agglutination does not occur, then the blood is considered Rh negative. When Rh-positive blood is transfused into a Rh-negative recipient, agglutinins to Rh + erythrocytes are gradually formed in the recipient's body over several months. Rh factor of erythrocytes.
The interaction of erythrocytes with anti-Rhesus serum is due to the presence in different areas membranes of several antigens (incomplete antigens). The most important of them are C, D, E, c, e. Antigen D has the most pronounced antigenic properties. For simplicity, blood containing D-erythrocytes is called Rh-positive (Rh + or Rh); and blood without such red blood cells is Rh-negative (Rh - or rh); 85% of Europeans have Rh+ blood; 15% - Rh -. The Rh+ phenotype may correspond to the DD or Dd genotype; and for the Rh phenotype - only the dd genotype.
One of the differences between the Rh and ABO systems, which has important practical significance, is that agglutinins of the ABO system are always contained in the human blood after the first months of life, while Rh agglutinins appear only after sensitization: contact of an Rh-negative individual with Rh -antigens. Therefore, the first transfusion of Rh-incompatible blood usually does not cause an obvious reaction. Antigen-antibody reactions appear only with repeated transfusion of such blood.
Another difference between the two systems is that most Rh agglutinins are incomplete immunoglobulin G antibodies, the size of which, unlike the size of complete ABO agglutinins, is small enough that they can penetrate the placental barrier.
Rh incompatibility and pregnancy. During pregnancy, small amounts of red blood cells can penetrate from the blood of an Rh-positive fetus into the blood of an Rh-negative mother. This leads to the production of agglutinins against Rh+ - red blood cells. Usually, only during childbirth a large amount (10-15 ml) of fetal red blood cells enters the mother’s blood. Since the antibody titer increases relatively slowly in the mother's blood (over several months), complications usually do not occur during the first pregnancy. However, during the second pregnancy of a Rh-negative woman with a Rh-positive fetus, the titer of antibodies in her blood can reach such a high level that as a result of the penetration of agglutinins through the placenta, the fetal red blood cells begin to be destroyed. This leads to serious disturbances in the functioning of the fetus and even to intrauterine death (erythroblastosis fetalis). The formation of antibodies in the body of a Rh-negative woman can be limited or completely suppressed through so-called anti-D prophylaxis. If immediately after childbirth (including premature birth) a woman is administered anti-D-globulin, then the Rh-positive red blood cells that have entered her blood will be destroyed and thereby the factor causing the production of antibodies will be eliminated immune system. Antigen-antibody reactions can also occur when mother and fetus are incompatible based on other group characteristics (in particular, ABO), but such reactions are usually mild.
Blood transfusion.
Blood transfusion rules:
1. Transfusion of isogroup blood.
2. Determination of group affiliation according to the ABO system.
3. Determination of Rh blood.
4. Determination of individual blood compatibility.
5. Conducting a biological test.
Determination of blood group according to the ABO system.
1. Method standard serums - standard sera of the first, second and third blood groups are used, which contain antibodies of the corresponding group: therefore, antigens are determined in the test blood.
2. Method standard red blood cells - standard red blood cells of the second and third groups are used, which contain the corresponding antigens (A, B); therefore, antibodies are determined in the blood being tested.
3. Method zoliclones - anti-A (alpha) coliclones are used and
anti-B (beta), these are monoclonal antibodies, we detect in the blood
antigens. Determination of Rh blood.
1. Method anti-Rhesus serums - Anti-Rh serum is used, which is layered on the wall of the test tube, followed by a drop of blood, the test tube is rocked for several minutes, then a drop of saline is added to eliminate false agglutination. If there is agglutination, it is Rh positive; in the absence of agglutination - Rh negative.
2. Method zoliclones - zolicone anti-D-cynep Determination of individual blood compatibility,
1. The donor's blood is mixed with the recipient's serum. If agglutination is present, the blood is incompatible; if absent - compatible.
2. The recipient's blood is mixed with the donor's serum. Grade
results are similar. Carrying out a biological test.
The first 45 ml of blood is transfused in fractions: after 3 minutes, 15 ml each and the patient’s condition is monitored. Incompatibility phenomena:
feeling of heat throughout the body, pallor skin, small and frequent pulse, decreased blood pressure; pain in the abdomen, head, lower back; urine is red-brown in color: “meat slop.”
This system is the main one that determines the compatibility or incompatibility of transfused blood. It includes two genetically determined important antigens: A and B - and two types of antibodies to them, agglutinins a and b. Combinations of agglutinogens and agglutinins determine 4 groups of the ABO system. This system is the only one where non-immune people have natural antibodies to the missing antigen in their plasma. Agglutinogen A in most people is well expressed (has great antigenic power): with anti-A antibodies (a), it gives a pronounced reaction of erythrocyte agglutination. In approximately 12% of individuals of groups A(11) and AB(IV), the antigen has weak antigenic properties; it is designated as A2 antigen. Thus, there is a group of antigens A: A1 (strong) and weaker A2, A3, A4, etc. The existence of weak A antigens should be remembered when determining blood groups, since red blood cells with such antigens are capable of giving only late and weak agglutination, which may lead to errors. Weak variants of the B antigen are very rare. Antibodies of the ABO system a (anti-A) and b (anti-B) are a normal property of blood plasma that does not qualitatively change during a person’s life, and and b are complete, cold antibodies. In most cases, they are not found in newborns and appear during the first three months of life or even a year. Group agglutinins reach full development by the age of 18, and in old age their titer (level) decreases, which is also observed in immunodeficiency states. In addition to the normally existing (natural) group antibodies a and b, in some cases immune antibodies anti-A and anti-B arise. Most common cause This is a pregnancy in which the mother and fetus have different blood groups, more often if the mother is group 1(0), the fetus is 11(A) or W(B). Determination of blood type is necessary for compatible blood transfusion. In this case, it is necessary to adhere to the rule: the donor's red blood cells should not contain an antigen corresponding to the recipient's antibodies, i.e. A and a, B and c, since otherwise there will be massive destruction of the injected red blood cells by the patient's antibodies - hemolysis, which can lead to the death of the recipient. Group antibodies of the donor can be ignored, since they are diluted by the recipient's plasma. Therefore, type O(I) blood, which does not contain agglutinogens, can be transfused to people of any blood group. Persons with blood type 0(1) are considered “universal donors”. Blood of group A(P) can be transfused into recipients of group A(P) and group AB(IV), which do not have agglutinins in the plasma. Blood of group B(III) can be transfused to persons with group B(III) and AB(IV).
Determination of blood groups of the ABO system is carried out using the following methods.
I. Determination of blood group using standard isohemagglutinating sera. With this method, the presence or absence of agglutinogens is determined in the blood and, based on this, a conclusion is made about the group affiliation of the blood being tested.
2. Determination of blood group using a cross method, i.e., simultaneously using standard isohemagglutinating sera and standard erythrocytes. With this method, as with the first, the presence or absence of agglutinogens is determined and, in addition, using standard red blood cells, the presence or absence of group agglutinins is determined.
3. Determination of blood group using monoclonal antibodies (COLICLONS).
ERRORS IN DETERMINING BLOOD TYPES
Technical errors. Violation of the stated rules for determining blood groups may lead to incorrect assessment of the reaction results. Deviations from the rules may include:
Use of substandard standard sera or red blood cells ( expired shelf life, contamination with them, drying of serums);
Mixing up blood samples;
Incorrect placement of standard sera or focytes in racks;
Incorrect order of applying standard reagents to the plate;
Incorrect serum to red blood cell ratio (not 10:1);
Study at a temperature of less than 15 ° C (cold agglutination occurs) or more than 25 ° C (agglutination slows down);
Failure to comply with the time required for the reaction (5 minutes);
Do not add saline solution followed by rocking the plate;
Do not use a control reaction with ABo(IV) group serum;
Use of dirty or wet pipettes, sticks, plates.
In all cases of unclear or questionable results, it is necessary to re-determine the blood group by cross-testing using standard sera from other series.
Errors associated with the biological characteristics of the blood being tested.
Incorrect identification of group A 2 and A 2 B. Red blood cells with weak antigen A form small, slowly appearing agglutinates with antiserum. The reaction can be taken into account as negative, i.e. group A 2 is mistakenly registered as O(1), and A 2 B as B(III). The risk of such an error is especially high in the simultaneous presence of technical errors (the ratio of serum to red blood cells is 10:1, the temperature is above 25 ° C, the results are taken into account earlier than 5 minutes).
Errors associated with the presence of nonspecific agglutinability of the studied erythrocytes. This phenomenon is observed in patients malignant tumors, leukemia, sepsis, burns, liver cirrhosis, autoimmune hemolytic anemia and is caused by dysproteinemia. Detects the presence of nonspecific agglutination by control with ABO (IV) group serum. In these cases, it is necessary to re-determine group membership using a cross-sectional method. In drops where agglutination is observed, you can add saline solution heated to 37°C. If necessary, you can wash the red blood cells being tested with warm (37°C) saline and re-determine the blood type.
Errors associated with the presence of extraagglutinins. In the blood serum of individuals of groups A2(P) and A2B(IV), antibodies to the A1 antigen - a1 - are found in approximately 1% of cases. This complicates the determination of blood group by the cross method, since the serum of such individuals agglutinates standard red blood cells of group A(P), i.e., manifests itself as serum of group 0(1).
In some diseases, a decrease in the agglutinability of erythrocytes, especially group A(P), is observed.
In immunodeficiency states, elderly people experience a decrease in the level of group agglutinins.
In all cases of obtaining a questionable result, the determination of blood group should be repeated using a cross-sectional method using sera of higher activity.
18. Antigens of the Rh system. Rh system groups. Clinical significance. Methods for determining Rh antigens and possible errors.
Rh antigens are the second most important in transfusion practice after ABO blood groups. During the period of active introduction of blood transfusions into the clinic, the number of post-transfusion complications after repeated transfusions of blood compatible with ABO antigens has increased significantly. The Rh system includes six antigens, to designate which two nomenclatures are used in parallel: Wiener (Rh 0, rh", rh", Hr 0, hr", hr"); Fischer and Reis (D, C, E, d, c, e).
Rh 0 - D, rh" - C, rh" - E, Hr 0 - d, hr" - c, hr" - e.
Since the Rho(D) antigen is the most active in this system, it is called the Rh factor. It is depending on the presence or absence of this factor that people are divided into Rh-positive (Rh+) and Rh-negative (Rh-). This division is accepted only in relation to recipients. Antigens rh"(C) and rh"(E) are less active than Rho(D), but antibodies to them can also be produced in people who do not contain antigens C and E in their red blood cells. Therefore, the requirements for red blood cells from Rh negative donors are more stringent. Red blood cells should not contain not only antigen D, but also C and E. Antigens Hro(d), hr"(c), hr"(e) are characterized by low activity, although hr"(c) antibodies can cause isoimmunological conflicts. 1-3% of Rh-positive individuals have a weak variant of the D - D antigen in their erythrocytes, which determines the presence of small, questionable agglutination when determining the Rh factor. In these cases, the Rh factor of the recipient's or pregnant woman's blood is indicated as Rh negative (Rh-), and the Rh factor of the donor's blood is indicated as Rh positive (Rh+). Transfusion of blood with antigen D to Rh-negative recipients is not allowed. Rh antigens are formed at 8-10 weeks of embryogenesis, and their antigenicity can even exceed the activity of antigens in adults. The Rh system, unlike the ABO system, does not have natural antibodies. Anti-Rhesus antibodies arise only after immunization of an Rh-negative organism as a result of a transfusion of Rh-positive blood or pregnancy with an Rh-positive fetus. In the body of sensitized individuals, antibodies to Rh antigens persist for several years, sometimes throughout life. In most cases, the titer of anti-Rhesus antibodies gradually decreases, but again increases sharply when Rh-positive blood enters the body again. Rh antibodies differ in specificity (anti-D, anti-III C, etc.) and in serological properties (complete and incomplete). Complete antibodies cause agglutination of red blood cells in a saline environment at room temperature. For agglutination to occur under the influence of incomplete antibodies, special conditions are required: elevated temperature, colloidal medium (gelatin, whey protein). Complete antibodies (IgM) are synthesized at the beginning of the immune reaction and soon disappear from the blood. Incomplete antibodies (IgG, IgA) appear later, take a long time to synthesize and cause the development of hemolytic disease in newborns, as they pass through the placenta and damage fetal cells.
Determination of Rh blood
The method for determining the Rh factor depends on the form of Rh antibodies in the standard serum and the method of its preparation. The anti-rhesus serum is accompanied by accompanying instructions describing the method for which this series of serum is intended.
For each study, a control must be placed to check the specificity and activity of anti-Rhesus serum. For control, standard Rh-positive erythrocytes of group 0(1) or the same group as the blood being tested are used, and standard Rh-negative erythrocytes must be of the same group as the blood being tested.
When determining Rh status by two series of standard sera in cases where they are used by different methods, the result is taken into account as true if it coincides in both series of studies after checking control samples confirming the specificity and activity of each series of anti-Rhesus serum, i.e. in the absence agglutination with standard Rh-negative erythrocytes of the same group and the presence of agglutination with standard Rh-positive erythrocytes of the same group or group 0(1) and in control samples without anti-Rhesus serum (reagent). If, when determining Rh status, a weak or questionable reaction is observed, then the person’s blood should be re-examined with the same and other series of anti-Rhesus serum and it is advisable to include serum containing complete antibodies. If all series of sera containing incomplete antibodies also give a weak or questionable reaction, and with complete antibodies the reaction is negative, this means that the red blood cells contain a weak type of rhesus antigen, the so-called factor D u. In these cases, the Rh-type of the patient's or pregnant woman's blood is indicated as Rh-negative (Rh-), and the Rh-type of the donor's blood as Rh-positive (Rh+), thus preventing the transfusion of his blood to Rh-negative recipients.
Determination of the Rh factor can also be carried out using the following methods.
Determination of the Rh factor Rh 0 (D) by a conglutination reaction using gelatin (in a test tube heated to 46-48 ° C).
Determination of the Rh factor Rho(D) by a conglutination reaction in a serum medium on a heated plane.
Determination of the Rh factor Rh 0 (D) by an agglutination reaction in a saline medium in small test tubes. The agglutination reaction in a saline environment is suitable only for working with serum containing complete Rh antibodies.
Determination of Rh factor Rh 0 (D) using monoclonal antibodies.
Determination of the Rh factor Rho(D) using an indirect Coombs test.
19 Anemia. Classification and a brief description of. Etiology and pathogenesis of anemia. Anemia (from the Greek anemia - lack of blood) is a large group of diseases that is characterized by a decrease in the amount of hemoglobin or hemoglobin and red blood cells per unit volume of blood. Anemia is different in etiology, development mechanisms, clinical and hematological picture, so there are many various classifications, but they are not perfect enough. L.I. Idelson proposed a working classification of anemia for clinicians: 1) acute posthemorrhagic anemia; 2) iron deficiency anemia; 3) anemia associated with impaired synthesis or utilization of porphyrins (sideroblastic); 4) anemia associated with impaired DNA and RNA synthesis (megaloblastic); 5) hemolytic anemia; 6) anemia associated with inhibition of proliferation of bone marrow cells (hypoplastic, aplastic); 7) anemia associated with the replacement of hematopoietic bone marrow by a tumor process (metaplastic).
Anemia can be like independent disease, so accompanying symptom or complication of certain internal diseases, infectious and oncological diseases. There are multifactorial anemias, that is, of mixed origin, for example: hemolytic anemia with iron deficiency, aplastic anemia with a hemolytic component, etc.
Depending on the:
1) the values of the color indicator distinguish between anemia:
Normochromic (color index 0.9-1.1);
Hypochromic (color index less than 0.85);
Hyperchromic (color index greater than 1.15);
2) the average diameter of red blood cells:
Normocytic (average erythrocyte diameter 7.2-7.5 µm)
Microcytic (the average diameter of red blood cells is less than 6.5 microns),
Macrocytic (the average diameter of red blood cells is more than 8.0 microns),
Megalocytic (the average diameter of erythrocytes is more than 12 microns);
3) the average volume of erythrocytes in femtoliters (fl, 1 fl is equal to 1 micron 3):
Normocytic (average erythrocyte volume 87±5 fL);
Microcytic (average red blood cell volume less than 80 fL);
Macrocytic (the average volume of red blood cells is more than 95 fL);
4) the level of reticulocytes in peripheral blood.
Regenerative (reticulocyte count 0.5-5%);
Hyperregenerative (the number of reticulocytes is more than 5%);
Hypo- and aregenerative (the number of reticulocytes is reduced or absent, despite severe anemia).
The reticulocyte level is an indicator of the regenerative function of the bone marrow in relation to erythropoiesis.
Normochromic anemias include acute posthemorrhagic (in the first days after blood loss), hypo- and aplastic, non-spherocytic hemolytic, autoimmune hemolytic, metaplastic (with leukemia, myeloma, etc.), as well as anemia that develops with endocrine disorders (hypofunction of the adrenal glands), kidney diseases, chronic infections.
Hypochromic anemias include iron deficiency, sideroblastic, some myelotoxic, and hemolytic (thalassemia).
B12-(folate)-deficiency and some hemolytic anemias are hyperchromic (hereditary microspherocytosis, if microspherocytes predominate among the red blood cells in the smear). Sometimes vitamin B1 2 -deficiency anemia can be normochromic.
Normocytic include acute posthemorrhagic, aplastic, autoimmune hemolytic anemia, etc.
Microcytic anemia includes iron deficiency and sideroblastic anemia, macrocytic anemia includes vigamin B12 (folate) deficiency anemia, etc.
Regenerative anemias include posthemorrhagic anemia; hyperregenerative - hemolytic anemia, especially the condition after a hemolytic crisis; hypo- and aregenerative - hypoplastic, aplastic anemia.
The bone marrow reacts to the development of iron deficiency, hemolytic anemia with irritation and red sprout hyperplasia. With hypoplastic anemia, there is a progressive decline in erythropoiesis up to its complete depletion.
20.Laboratory diagnostics iron-saturated and iron-unsaturated anemias. Iron-deficiency anemia. Types of iron deficiency. Laboratory tests reflecting iron deficiency in the body. Picture of peripheral blood and bone marrow in IDA. Laboratory diagnosis of sideroblastic anemia. Metabolism and role of iron in the body
Iron is of great importance for the body; it is part of hemoglobin, myoglobin, and respiratory enzymes. It is distributed among fixed assets.
Hemoglobin fund. Hemoglobin iron makes up 60-65% of the total iron content in the body.
Emergency Fund. This is the iron of ferritin and hemosiderin, which are deposited in the liver, spleen, bone marrow, and muscles. Makes up 30-40% of the iron level in the body. Ferritin is a water-soluble complex of ferric iron and apoferritin protein, containing 20% iron. It is a labile fraction of the iron reserve fund. If necessary, it is easily used for the needs of erythropoiesis. Hemosiderin is a water-insoluble protein, similar in composition to ferritin, but contains a larger amount of iron - 25-30%. It is a stable, firmly fixed fraction of iron reserves in the body.
The transport fund is represented by iron bound to the transport protein transferrin. Constitutes 1% of the iron content in the body.
The tissue fund is represented by iron, iron-containing enzymes (cytochromes, peroxidase, etc.), myoglobin. Constitutes 1% of the iron content in the body.
The total iron content in the body of adults is 4-5 g. It enters the body with the diet. Contained in animal products and plant origin(meat, especially beef, liver, eggs, legumes, apples, dried apricots, etc.). Iron is absorbed much better from animal products than from plant products, since it is contained in them in the form of heme. Thus, 20-25% is absorbed from meat, 11% from fish, plant products- 3-5% of the iron they contain. Iron absorption is promoted ascorbic acid, organic acids (citric, malic, etc.), inhibit the absorption of tannin, high fat content in the diet. Absorption of iron from food products limited. 2-2.5 mg of iron is absorbed per day; for a short time after severe bleeding, up to 3 mg of iron can be absorbed. The main amount of iron is absorbed in the duodenum and in the initial part jejunum. A small amount of iron can be absorbed in all parts of the small intestine.
Iron absorption occurs in two stages: 1) the intestinal mucosa captures iron supplied with the diet; 2) iron from the intestinal mucosa passes into the blood, is loaded onto transferrin and delivered to places of use and depot. Transferrin also transfers iron from its funds and cells of the phagocytic mononuclear system, in which destruction of red blood cells occurs, to the bone marrow, where it is partially used for the synthesis of hemoglobin, and partially deposited in the form of iron reserves, as well as to other iron storage sites. Typically, 1/3 of transferrin is bound to iron. It is called bound transferrin or serum iron. Normal serum iron levels in men and women are 13-30 and 12-25 µmol/l, respectively. The portion of transferrin not bound to iron is called free transferrin or unsaturated, latent serum iron-binding capacity. The maximum amount of iron that transferrin could attach before its saturation is designated as the total iron-binding capacity of serum (TIBC) (normally 30-85 µmol/l). The difference between TIBC and serum iron values reflects the latent iron-binding capacity, and the ratio of serum iron to TIBC, expressed as a percentage, reflects the percentage of transferrin saturation with iron (normal 16-50%). To judge the amount of iron reserves in the body, the following is carried out:
Study of serum ferritin levels using radioimmunoassay methods;
Desferal test. Desferal (desferoxamine) is a complexone that, after being introduced into the body, selectively binds to iron reserves, i.e., ferritin iron, and removes it in the urine. The patient is injected intramuscularly with 500 mg of desferal once, daily urine is collected and the iron content in it is determined. After administration of desferal, 0.8 to 1.2 mg of iron is normally excreted in the urine, while in patients with iron deficiency anemia or in the presence of hidden iron deficiency, the amount of iron excreted in the urine decreases sharply;
Counting the number of sideroblasts in the bone marrow puncture, and siderocytes in the peripheral blood. Sideroblasts are normoblasts, i.e., nucleated cells of the red row, in the cytoplasm of which blue granules of iron reserves - ferritin - are detected. Normally, 20-40% of normoblasts are sideroblasts. Siderocytes are red blood cells in which ferritin granules are found. Normally in the peripheral blood: up to 1% siderocytes. Ferritin granules in sideroblasts and siderocytes are revealed by special staining with Prussian blue.
The body is characterized by physiological losses of iron in urine, feces, bile, exfoliated cells of the intestinal mucosa, sweat, and when cutting hair and nails. Women lose iron with their periods.
The development of iron deficiency anemia is preceded by hidden (latent) iron deficiency. Patients have complaints and Clinical signs, characteristic of iron deficiency pyaemia, but less pronounced (weakness, moderate pallor of the skin and visible mucous membranes, headaches, palpitations, often perversion of taste and smell, dry skin, brittle nails, etc.). The examination does not yet reveal changes in the content of hemoglobin, red blood cells and other indicators of peripheral blood. But disturbances in iron metabolism are revealed: serum iron decreases, total and latent iron-binding abilities of serum increase, the percentage of transferrin saturation decreases, and the level of iron reserves decreases. This is sideropenia without anemia. Hidden iron deficiency can develop at any age, and women, adolescents and children are especially likely to suffer from it. If hidden iron deficiency is not compensated, but deepens, iron deficiency anemia develops.