COMPATIBILITY TESTING

The term compatibility testing refers to the set of procedures required before blood can be
issued as being safe for transfusion. By ensuring that there are no antibodies in the patient's
serum that react with the red cells for transfusion, the donor's blood should not cause any
adverse reactions and the red cells will have a maximum survival time following transfusion.
Blood compatibility testing is conducted in a medical laboratory to identify potential
incompatibility between individuals' and blood donor's blood types, which can occur
in blood transfusion. It is also used to diagnose and prevent some complications of
pregnancy that can occur when the baby has a different blood group from the
mother.
Blood compatibility testing generally makes use of reactions between blood group
antigens and antibodies specifically the ability of antibodies to cause red blood cells
to clump together when they bind to antigens on the cell surface, a phenomenon
called agglutination. Techniques that rely on antigen-antibody reactions are
termed serologic methods, and several such methods are available, ranging from
manual testing using test tubes or slides to fully automated systems. Blood types can
also be determined through genetic testing, which is used when conditions that
interfere with serologic testing are present or when a high degree of accuracy in
antigen identification is required.
Compatibility testing involves:
1. Checking the patient’s records for the results of: previous blood grouping, the
presence of any antibodies , details of past transfusions, the reason for
transfusion.
2. This information should be given on the blood request form, but it is worth also
checking your own laboratory records.
3. Performing an ABO and RhD group on the patient’s blood sample and checking to
ensure that these match any previous results you have.
4. Performing an antibody screen, if possible.
Performing the final compatibility test, the cross-match, cross-match which is the test
between the patient’s serum and the donor red cells to detect any antibodies in the
patient's serum that react with the donor red cells. This is sometimes referred to as the
major cross-match. The minor cross match is the testing of the patient’s red cells with donor
serum to detect the presence of any antibodies in the donor’s serum that might react with
the patient’s red cells. In the majority of cases, the minor cross-match is no longer required
since the donor’s serum is checked for the presence of antibodies when it is grouped.
Blood group systems other than ABO and Rh
Other than the ABO and Rh blood group systems, there are some 20 blood group systems,
such as Duffy, Kidd and Kell, on human red cells.
These might lead to the production of antibodies if a person lacking one of these antigens is
sensitized, by pregnancy or transfusion, to that antigen. If these antibodies are not detected
in the compatibility test, they could cause severe transfusions reactions.
Crossmatching in compatibility test:
Crossmatching, which is routinely performed before a blood transfusion, involves adding the
recipient's blood plasma to a sample of the donor's red blood cells. If the blood is
incompatible, the antibodies in the recipient's plasma will bind to antigens on the donor red
blood cells. This antibody-antigen reaction can be detected through visible clumping or
destruction of the red blood cells, or by reaction with anti-human globulin, after
centrifugation.
If the transfusion recipient has a negative antibody screen and no history of antibodies, an
"immediate spin" crossmatch is often performed: the red blood cells and plasma are
centrifuged immediately after mixing as a final check for incompatibility between ABO blood
types. If an clinically significant antibody is detected (or was in the past), or if the immediate
spin crossmatch demonstrates incompatibility, a "full" or "IgG crossmatch" is performed,
which uses the indirect antiglobulin test to detect blood group incompatibility caused by IgG
antibodies. The IgG crossmatching procedure is more lengthy than the immediate spin
crossmatch, and in some cases may take more than two hours.

Individuals who have a negative antibody screen and no history of antibodies may also
undergo an "electronic crossmatch", provided that their ABO and Rh type has been
determined from the current blood sample and that the results of another ABO/Rh type are
on record. In this case, the recipient's blood type is simply compared against that of the
donor blood, without any need for serologic testing. In emergencies, blood may be issued
before crossmatching is complete.
Compatibility test of ABO:
When selecting blood for transfusion, it is important that blood of the correct ABO group is
used. Red cells that are incompatible with the ABO antibodies in the patient’s plasma can
cause fatal haemolytic reactions.
Red cell components:
In red cell transfusion, there must be ABO and Rh compatibility between the donor’s red
cells and the recipient’s plasma.
1. Group O individuals can receive blood from group O donors only.
2. Group A individuals can receive blood from group A and O donors.
3. Group B individuals can receive blood from group B and O donors.
4. Group AB individuals can receive blood from AB donors, and also from group A, B
and O donor.
Plasma and components containing plasma:
In plasma transfusion, group AB plasma can be given to a patient of any ABO group because
it contains neither anti-A nor anti-B antibody
1. Group AB plasma (no antibodies) can be given to any ABO group.
2. Group A plasma (anti-B) can be given to group O and A patients.
3. Group B plasma (anti-A) can be given to group O and B patients.
4. Group O plasma (anti-A + anti-B) can be given to group O patients only.
Method:
Blood typing can be performed using test tubes, microplates, or blood typing slides. The
tube method involves mixing a suspension of red blood cells with antisera (or plasma, for
reverse grouping) in a test tube. The mixture is centrifuged to separate the cells from the
reagent, and then resuspended by gently agitating the tube. If the antigen of interest is
present, the red blood cells agglutinate, forming a solid clump in the tube. If it is absent, the
red blood cells go back into suspension when mixed.
The microplate method is similar to the tube method, except rather than using individual
test tubes, blood typing is carried out in a plate containing dozens of wells, allowing multiple
tests to be performed at the same time. The agglutination reactions are read after the plate
is centrifuged.
Antibody screening and identification can also be carried out by the tube method. In this
procedure, the plasma and red cells are mixed together in a tube containing a medium that
enhances agglutination reactions, such as low ionic strength saline (LISS). The tubes are
incubated at body temperature for a defined period of time, then centrifuged and examined
for agglutination or hemolysis; first immediately following the incubation period, and then
after washing and addition of anti-human globulin reagent.
Crossmatching, likewise, may be performed by the tube method; the reactions are read
immediately after centrifugation in the immediate spin crossmatch, or after incubation and
addition of AHG in the full crossmatching procedure.
The slide method for blood typing involves mixing a drop of blood with a drop of antisera on
a slide. The slide is tilted to mix the cells and reagents together and then observed for
agglutination, which indicates a positive result. This method is typically used in under-
resourced areas or emergency situations; otherwise, alternative methods are preferred.
Genotyping:
 Genetic testing can be used to determine a person's blood type in certain situations
where serologic testing is insufficient. For example, if a person has been transfused
with large volumes of donor blood, the results of serologic testing will reflect the
antigens on the donor cells and not the person's actual blood type.Individuals who
produce antibodies against their own red blood cells or who are treated with certain
drugs may show spurious agglutination reactions in serologic testing, so genotyping
may be necessary to determine their blood type accurately. Genetic testing is
 required for typing red blood cell antigens for which no commercial antisera are
available.
 The AABB recommends RhD antigen genotyping for women with serologic weak D
phenotypes who have the potential to bear children. This is because some people
with weak D phenotypes can produce antibodies against the RhD antigen, which can
cause hemolytic disease of the newborn, while others cannot. Genotyping can
identify the specific type of weak D antigen, which determines the potential for the
person to produce antibodies, thus avoiding unnecessary treatment with Rho(D)
immune globulin.
 Genotyping is preferred to serologic testing for people with sickle cell disease,
because it is more accurate for certain antigens and can identify antigens that cannot
be detected by serologic methods.
 Genotyping is also used in prenatal testing for hemolytic disease of the newborn.
When a pregnant woman has a blood group antibody that can cause HDN, the fetus
can be typed for the relevant antigen to determine if it is at risk of developing the
disease. Because it is impractical to draw blood from the fetus, the blood type is
determined using an amniocentesis sample or cell-free fetal DNA isolated from the
mother's blood.
 The father may also be genotyped to predict the risk of hemolytic disease of the
newborn, because if the father is homozygous for the relevant antigen (meaning
having two copies of the gene) the baby will be positive for the antigen and thus at
risk of developing the disease. If the father is heterozygous (having only one copy),
the baby only has a 50% chance of being positive for the antigen.

ABO and Rh typing:

In ABO and Rh typing, reagents containing antibodies against the A, B, and RhD antigens are
added to suspensions of blood cells. If the relevant antigen is present, the antibodies in the
reagent will cause the red blood cells to agglutinate (clump together), which can be
identified visually.
In addition to identifying the ABO antigens, which is termed forward grouping, routine ABO
blood typing also includes identification of the ABO antibodies in the person's plasma. This is
called reverse grouping and it is done to confirm the ABO blood type. In reverse grouping,
the person's plasma is added to type A1 and type B red blood cells.
The plasma should agglutinate the cells that express antigens that the person lacks, while
failing to agglutinate cells that express the same antigens as the patient. If this does not
occur, further testing is required.
Agglutination is scored from 1+ to 4+ based on the strength of the reaction. In ABO typing, a
score of 3+ or 4+ indicates a positive reaction, while a score of 1+ or 2+ is inconclusive and
requires further investigation.
Serological methods:
  Serologic methods for blood compatibility testing make use of these antibody-
antigen reactions. In blood typing, reagents containing blood group antibodies,
called antisera, are added to suspensions of blood cells. If the relevant antigen is
present, the antibodies in the reagent will cause the red blood cells to agglutinate
(clump together), which can be identified visually.In antibody screening, the
individual's plasma is tested against a set of red blood cells with known antigen
profiles; if the plasma agglutinates one of the red blood cells in the panel, this
indicates that the individual has an antibody against one of the antigens present on
the cells. In crossmatching, a prospective transfusion recipient's plasma is added to
the donor red blood cells and observed for agglutination (or hemolysis) to detect
antibodies that could cause transfusion reactions.
 Blood group antibodies occur in two major forms: immunoglobulin M (IgM) and
immunoglobulin G (IgG). Antibodies that are predominantly IgM, such as the ABO
antibodies, typically cause immediate agglutination of red blood cells at room
temperature. Therefore, a person's ABO blood type can be determined by simply
adding the red blood cells to the reagent and centrifuging or mixing the sample and
in crossmatching, incompatibility between ABO types can be detected immediately
after centrifugation.
 RhD typing also typically uses IgM reagents although anti-RhD usually occurs as IgG
in the body. Antibodies that are predominantly IgG, such as those directed towards
antigens of the Duffy and Kidd systems, generally do not cause immediate
agglutination because the small size of the IgG antibody prevents formation of a
lattice structure. Therefore, blood typing using IgG antisera and detection of IgG
antibodies requires use of the indirect antiglobulin test to demonstrate IgG bound to
red blood cells.
 In the indirect antiglobulin test, the mixture of antiserum or plasma and red blood
cells is incubated at 37 °C (99 °F), the ideal temperature for reactivity of IgG
antibodies. After incubation, the red blood cells are washed with saline to remove
unbound antibodies, and anti-human globulin reagent is added. If IgG antibodies
have bound to antigens on the cell surface, anti-human globulin will bind to those
antibodies, causing the red blood cells to agglutinate after centrifugation.
 If the reaction is negative, "check cells"—reagent cells coated with IgG—are added
to ensure that the test is working correctly. If the test result is indeed negative, the
check cells should react with the unbound anti-human globulin and demonstrate
agglutination.
Medical use of compatibility test:
Blood compatibility testing is routinely performed before a blood transfusion. The full
compatibility testing process involves ABO and RhD (Rh factor) typing; screening for
antibodies against other blood group systems; and crossmatching, which involves testing
the recipient's blood plasma against the donor's red blood cells as a final check for
incompatibility. If an unexpected blood group antibody is detected, further testing is
warranted to identify the antibody and ensure that the donor blood is negative for the
relevant antigen.
Serologic crossmatching may be omitted if the recipient's antibody screen is negative, there
is no history of clinically significant antibodies, and their ABO/Rh type has been confirmed
against historical records or against a second blood sample; and in emergencies, blood may
be transfused before any compatibility testing results are available.
Blood compatibility testing is also routinely performed on pregnant women and on the cord
blood from newborn babies, because incompatibility puts the baby at risk for developing
hemolytic disease of the newborn. It is also used before hematopoietic stem cell
transplantation, because blood group incompatibility can be responsible for some cases of
acute graft-versus-host disease.
Transfusion reactions occurs:
Fortunately, the majority of blood transfusions take place without any adverse effects on
the patient. Occasionally, however, patients will react to transfused blood even though the
laboratory tests carried out before transfusion showed the blood to be compatible.
The severity of reaction that a patient suffers can vary from a mild reaction, which leads to
little more than a headache with a slight rise in body temperature, to the more severe
haemolytic form which, in rare cases, can be fatal.
Transfusion reactions fall mainly into three categories:
1. febrile reactions
2. allergic reactions
3. haemolytic reactions.
Febrile reactions:
Febrile reactions lead to headache followed by a sudden chill, shivering and a rise in body
temperature. These reactions are rarely severe and respond rapidly to medical treatment.
Allergic reactions:
Severe allergic reactions, sometimes called anaphylactic reactions are comparatively rare. In
such cases, the patient can suffer urticaria of the skin, moderate bronchial spasm and
possible laryngeal oedema. Reactions of this kind are rare and respond rapidly to medical
treatment.
Haemolytic reactions:
Haemolytic reactions are the most severe of the three types of transfusion reaction and are
initiated by:

 Antibody in the patient’s serum reacting with its corresponding antigen on the donor
red cells.
 Antibody in the donor plasma reacting with its corresponding antigen on the
patient’s red cells.
laryngeal oedema:
Swelling of the larynx creating difficulty in breathing.
Haemoglobin-aemia:
Free haemoglobin in the bloodstream (plasma).
Macrophage:
A phagocytic cell type found in the bloodstream as well as tissues. Macrophages ingest
bacteria and cell debris.