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ISBT Science Series (2008) 3, 68–92

© 2008 The Authors.


SECTION 6 Journal compilation © 2008 Blackwell Publishing Ltd.

Blood group systems


Blackwell Publishing Ltd

E. Smart & B. Armstrong

– well known antigens


Introduction
– antigen frequencies
This section will cover the major blood group systems, – antibody characteristics
some of the other blood group systems and will also include – significance in transfusion
information on HLA and notes on platelet antigens. Although – significance in haemolytic disease of the fetus and
some references will be made to the molecular structures, the newborn
detailed molecular structures and recent advances in DNA – features and practical application
technology are not within the scope of this publication. • MNS
• P
• Kell
Learning objectives
• Duffy
By the end of the section, the student should be able to • Kidd
describe the following in simple terms: • Lewis
• Blood group terminology • Lutheran
• Functions of blood groups • I blood group system
• ABO and H blood group systems • Additional blood group systems/collections/antibodies
– ABO grouping reacting with high and low frequency antigens
– inheritance of ABO blood groups • Polyagglutination
– ABO blood group frequencies • Human leucocyte antigen system
– production of ABO antigens – disease association
– H-deficient phenotypes and Bombay Oh phenotype – transplantation
• Subgroups within the ABO system – transfusion
• ABO system antibodies – pregnancy
• Clinical significance of ABO system: – parentage testing
– clinical significance in transfusion • Human platelet antigen systems
– clinical significance in haemolytic disease of the fetus • Fetomaternal alloimmune thrombocytopaenia.
and newborn
• Lectins (plant agglutinins)
• ABH secretion
Blood group terminology
• Unique features of the ABO system Currently, 29 different blood group systems are known, nine
• Rh blood group system of which are considered to be the major blood group systems.
• Rh genetics and inheritance In addition there are various blood group antigens that
– molecular studies have been allocated to collections, low incidence antigens
– terminology (700 Series) or high incidence antigens (900 Series) according
– frequencies to the International Society of Blood Transfusion (ISBT)
• Rh typing Committee on Terminology for the Red Cell Surface Antigens,
• Rh antigens which is the committee responsible for the terminology and
• Clinical significance of Rh system allocation of antigens to the appropriate system.
– clinical significance in transfusion There are strict criteria for:
– clinical significance in haemolytic disease of the fetus • The allocation of blood group antigens to a new blood
and newborn group system (the antigen must be shown to be an inherited
• Unique features of the Rh system character defined by a human alloantibody, the gene
• Other major blood group systems (MNS, P, Kell, Duffy, encoding it must have been identified and sequenced, and
Kidd, Lewis, Lutheran): the chromosomal location must be known).
– date of discovery • The allocation of a new specificity to an existing system.

68
Introduction to Blood Transfusion Technology 69

• The establishment of a collection (requires two or more Table 6·2 Other blood group systems
antigens that are related serologically, biochemically or
genetically, but do not fulfil the requirements for a blood Blood group Chromosome
group system). ISBT No system name Main antigens location no.
• Inclusion into the 700 series (incidence of less than 1% of
010 Diego Dia, Dib, Wra, Wrb 17
the population and distinct from other systems and
011 Yt Yta, Ytb 7
collections).
012 Xg Xga X
• Inclusion into the 900 series (incidence of > 90% in most 013 Scianna Sc1, Sc2 1
populations tested and distinct from other high frequency 014 Dombrock Doa, Dob, Gya, Hy, Joa 12
antigens). 015 Colton Coa, Cob, Co3 7
Various terminologies have been used to describe the 016 Landsteiner-Weiner LW 19
different blood group systems and their antigens and respec- 017 Chido/Rodgers CH/RG 6
tive antibodies ever since the ABO blood group system was 018 H H 19
first discovered in 1900. In 1980 an ISBT committee was 019 Kx Kx X
tasked to devise a genetically based numerical terminology 020 Gerbich Ge2, Ge3, Ge4 2
021 Cromer Cra 1
for red cell antigens. This is an ongoing process and new
022 Knops Kna, Knb 1
information regarding the antigens and apparent new
023 Indian Ina, Inb 11
antigens are reviewed by the committee on a regular basis.
024 Ok Oka 19
The numerical terminology was primarily designed to 025 Ralph MER2 11
facilitate computer input. The alternative/popular terminologies 026 John Milton Hagan JMH 15
are commonly used, both in everyday communication, in the 027 I I 6
laboratories and in publications. 028 Globoside P 3
Note: The term group or type can be used interchangeably 029 Gill GIL 9
when discussing blood grouping or typing. Further notes on
Rh terminology will be found in the Rh section.
The number of antigens per blood group system, collection,
and series varies tremendously from 1 in the P system to more information is available particularly since the development
than 30 in the Rh and MNS systems. of molecular genetic techniques. Less is known about the
Table 6·1 shows the major blood group systems. Note that actual function of the blood groups. The red cell is a complex
H antigen is in a separate system, H Blood Group System 018, structure, and the red cell membrane contains many surface
and is not part of the ABO system. proteins that are anchored to the membrane or cross the
Table 6·2 gives information on the blood group systems lipid bilayer one or more times. Many of the proteins on the
other than the nine major systems. surface of the red cells are polymorphic and carry the different
blood groups. The functions of some of the red cell membrane
proteins have been identified, and other functions have been
Functions of blood groups deduced from the structures of the protein. Studies on the null
The structures of the different blood group systems and their phenotypes which occur in most blood group systems have
antigens have been studied extensively, and a wealth of contributed to the information. The ABO, H, I, P1 and P blood
groups are carbohydrate structures on the red cell membrane
glycolipids and glycoproteins and less is known about their
Table 6·1 Major blood group systems (9 of 29)
function. Table 6·3 provides a list of the functions of the
Blood group Chromosome blood groups.
ISBT No system name Major antigens location no. The format for describing the blood group systems in this
publication is as follows:
001 ABO A, B, A1B, A1 9 Antigen frequencies are given as approximate percentages,
002 MNS M, N, S, s, U 4 simply to make them easier to remember. This will sometimes
003 P P1 22 result in the total being slightly more or less than 100%. The
004 Rh D, C, E, c, e 1 figures for black people apply to published data, or to surveys
005 Lutheran Lua, Lub 19
performed in southern Africa.
006 Kell K, k, Kpa, Kpb, Jsa, Jsb 7
• When an antibody is stated to cause haemolytic disease of
007 Lewis Lea, Leb 19
the fetus and newborn (HDFN) and when it has an optimum
008 Duffy Fya, Fyb, Fy3 1
009 Kidd Jka, Jkb, Jk3 18
reaction temperature of +37°C using the IAT technique, it
is presumed to be a type IgG antibody.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
70 Blood group systems

Table 6·3 Functions of blood groups Table 6·4 ABO groups

Blood groups Red cell function ABO Group Antigens on red cells Antibodies in serum/plasma

Rh, Kidd, Kx, Diego, Colton, Gill Membrane transporters A A Anti-B


Kell, Yt Membrane bound enzymes B B Anti-A
MNS, Gerbich Structural proteins O None Anti-A,B
Duffy Chemokine receptor AB A and B None
Lutheran, LW, Xg, Indian Cell adhesion molecules
Cromer, Knops Complement regulation
ABO, H, I, P1 and P Carbohydrate structures which for the majority of serious and/or fatal transfusion reactions
contribute to the glycocalyx and are usually caused by technical, clerical or administra-
tive errors.

• When an antibody is described as a saline agglutinin and


ABO grouping
when it reacts best at cold temperatures, it is presumed to
be a type IgM antibody. The ABO system is unique in that whenever the A or B
antigens are not present on the red cells, the corresponding
The two most clinically significant blood group systems antibody is present in the serum/plasma. Anti-A and anti-B
are the ABO and the Rh blood group systems. alloagglutinins are therefore often referred to as being
‘naturally occurring’.
ABO grouping can therefore be performed by:
ABO and H blood group systems • Typing the red cells for the presence or absence of the A
Although the ABO and H are two different blood group systems and/or B antigens. This is known as forward grouping.
genetically they will be described together as they are closely • Testing the serum/plasma for the presence or absence of
related, both at the biochemical and phenotype level. anti-A and/or anti-B antibodies. This is known as reverse
The ABO system is the most important blood group system grouping.
in transfusion therapy and was the first blood group system • The forward and reverse grouping results should correlate;
to be discovered. This great contribution to medicine was refer to Landsteiner’s rule.
made by Landsteiner in 1900 when he observed that ‘the The general population can then be divided into four ABO
serum of healthy humans not only has an agglutinating groups as shown in Table 6·4, based on the forward and
effect on animal blood corpuscles, but also on human blood reverse grouping.
corpuscles from different individuals’. The following year, in It should be noted that the anti-A,B produced by a group
1901, Landsteiner was able to recognize two antigens on the O individual is different from anti-A+B, which is a mixture
red cells by separating and mixing the cells and sera of several of anti-A from one source and anti-B from another source.
individuals. He called the antigens A and B. Those individuals Anti-A,B detected in group O individuals is an antibody that
with the A antigen on their red cells were called group A; will react with group A and group B cells. The monoclonal
those with the B antigen, group B. Many individuals lack the anti-A,B reagents available commercially will detect the
A and the B antigens and were termed group C, which was weak A group Ax.
later termed group O. The least common group, called AB,
was found by several of Landsteiner’s students in 1902.
Inheritance of the ABO blood groups
Group AB individuals have both A and B antigens on their
red cells. Landsteiner found that the serum of an individual The ABO genes are located on chromosome number 9. The
always contained antibodies to the antigen, which was not inheritance in the ABO system is controlled by various
present on that individual’s red cells. Thus, group A individuals alleles: A1, A2, B and O and a series of rare alleles A3, Ax and
will have anti-B antibodies in their serum and group B Am (etc). The O allele (which does not produce an antigenic
individuals will have anti-A antibodies in their serum. These product) is recessive to the A and B alleles, which are co-
facts became known as Landsteiner’s Rule which states, dominant. The ABO phenotype is shown by ABO grouping
‘(In the ABO system) the antibody to the antigen lacking on laboratory tests on a blood specimen but this is not necessar-
the red cells is always present in the serum or plasma.’ ily the genotype of the individual. For example, blood of
The regular presence of anti-A and/or anti-B antibodies phenotype A1 can represent one of several possible genotypes:
means that it is critical for patient safety and good transfu- A1A1, A1A2, A1A3, A1Ax, A1Am (etc) or A1O.
sion practice that ABO groups are performed, recorded and Although each individual has two ABO genes, serological
interpreted correctly. ABO incompatibilities are responsible tests do not reveal the O allele in the A and B phenotypes, nor

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 71

Table 6·5 ABO group/phenotype and possible genotypes (simplified) Table 6·6 Example of ABO blood group distribution in percentage – one
survey in southern Africa
Blood group/phenotype Possible genotypes
Group White Black Indian
A1 A1A1, A1A2, A1A3, A1Ax, A1Am, A1O
A2 A2A2, A2A3, A2Ax, A2Am, A2O A 39 30 24
B BB, BO B 10 19 32
AB A1B, A2B, A3B, AxB, AmB O 47 46 37
O OO AB 4 5 7

can an allele producing a weak form of A be recognized if an precursor substrate, which is a carbohydrate chain already
allele higher in the scale of A antigen production is simulta- present on the red cell membrane. Once this has been performed,
neously present. The genotype can however be determined by the A- and B-transferases can act. The A-transferase adds
DNA analysis of the gene or may be determined by family another sugar called N-acetyl-D-galactosamine, which results
studies. Table 6·5 shows ABO blood group/phenotype with in the expression of A antigen on the red cells. Similarly, the
possible genotypes (simplified), including some of the rare B-transferase adds the sugar D-galactose and the cells then
alleles. also express B antigen. These red cells type as group AB.
Group A antigen is expressed when H and A transferases
are the two enzymes present; group B antigen is expressed
ABO blood group frequencies
when the H and B transferases are the enzymes present, and
Many populations have been studied world-wide, and it has in group O only H transferase is present. Figure 6·1 shows a
been shown that the frequency of the ABO blood group genes simplified diagram to indicate the structural differences in
varies between different populations. Note the variation the molecules that result in ABH antigen expression.
shown in Table 6·6 as an example of ABO blood group The expression of A, B or AB antigens results in a relative
distribution. masking of the H antigen. Thus, A1, B or A1B cells express
only small quantities of H.
The A2 allele is less effective than the A1 allele in masking
Production of ABO antigens
the H determinant, and A2 cells therefore have considerably
The ABO red cell antigens expressed on the red cells are more H antigen and less A antigen than do A1 cells. The O allele
dependent on the presence of both the H gene, and the ABO in double dose leads to the expression of H specificity alone,
genes. The loci for the H and ABO genes are not linked resulting in group O individuals having abundant H antigen.
although they are related and they are therefore two separate The amount of H antigen that is present in red cells of
blood group systems. The H, A and B genes do not code different groups, from left or right in decreasing order, is as
directly for red cell antigens, but for enzymes known as follows: most H antigen: O → Weak A → A2 → A2B → B →
transferases. The H-transferase adds the sugar L-fucose to a A1 → A1B → least H antigen.

Fig. 6·1 Simplified diagram to indicate


structural differences in ABH antigen
composition.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
72 Blood group systems

The A, B and H antigens are detectable long before birth.


Subgroups within the ABO system
The ABH antigen strength usually peaks at between 2 and
4 years of age and then remains relatively constant in most
Subgroups of A
individuals. It may not be possible to distinguish between
group A1 and A2 groups at birth as the antigens may not yet About 10 years after the discovery of the ABO groups, the
be fully expressed. subgroups of A were described. It was observed that not all
group A bloods tested gave similar results with anti-A. Those
unusual bloods which gave weaker reactions became known
H-deficient phenotypes
as ‘weak A’. Furthermore, it was realized that the common A
Although the ABO and H are two different blood group antigen occurred in two forms: A1 and A2. Later studies on
systems genetically (H Blood Group System: Number 018), transferase enzymes of group A1 and group A2 individuals
they are closely related at the biochemical and phenotype showed that less antigenic sites are produced in group A2
level. The H-deficient phenotypes are very rare and include individuals as the enzyme is less effective in converting the
a total deficiency in H antigen (Bombay or Oh phenotype) or precursor H substance into A antigen. However, with the use
a partial deficiency (Parabombay). of monoclonal anti-A blood grouping reagents, little if any
difference between the reactions of group A1 and group A2
cells can be detected in the laboratory. About 99.9% of all
Bombay Oh phenotype
group A bloods from white people and about 96% of group
The Bombay or Oh phenotype, in which the cells lack the H A bloods from black people, in one survey, were either group
antigen, arises when the individual has not inherited the very A1 or A2, with group A1 being more frequent than group A2
common gene H. As there is no H gene present, the H- in both populations. A higher incidence of ‘weak’ A was
transferase enzyme is absent. The precursor substance remains detected in the black people.
unchanged and no molecules of L-fucose are present on the The anti-A found in the serum/plasma of group B individuals
precursor substrate in the red cell membrane. The individual consists of two separate antibody specificities, anti-A and
may have inherited the A and/or B genes, which code nor- anti-A1, the latter being specific for the A1 type. Group A or
mally for the appropriate transferases. However, without the AB individuals who lack the A1 component may form an
single terminal carbohydrate (sugar) L-fucose at the end of irregular, cold reacting anti-A1 antibody in their serum. The
the substrate protein, these transferases are non-reactive. The lectin Dolichos biflorus or monoclonal anti-A1 reagents, are
Bombay Oh phenotype therefore results when the individual usually used to type red cells for the A1 antigen.
has inherited a double dose of a rare recessive allele, known
as h. The gene h does not code for H transferase. Individuals
Further subgroups of group A and subgroups of
who have inherited HH or Hh genes produce normal amounts
group B
of H transferase.
Bombay Oh individuals are extremely rare. Those who were A number of further subgroups of group A and subgroups of
originally shown to carry the trait were Indians whose ancestors group B also occur. The subgroups are caused by genetic
originated in Bombay, hence the name Bombay Oh. Their cells variations that result in a variety of weakened expressions of
are not agglutinated by anti-A, -B, -A,B or -H. Bombay Oh the antigens. The subgroups cannot be detected when inherited
individuals have powerful anti-H, -A and -B antibodies in with a normal A or B gene. The subgroups may be detected
their serum. They should therefore be transfused only with in the laboratory when weak or unexpected negative results
type Oh blood. Table 6·7 shows the difference between blood are obtained with the forward grouping and/or anomalous
group O and blood group Oh. results with the reverse grouping.

Weak A
The term weak A covers a large range of reactivity, some
bloods giving clear (although weak) results and other bloods
Table 6·7 Differences in blood groups O and Oh
giving such weak reactions that detection may prove difficult.
Forward grouping Reverse grouping The weak A types include A3, Am, Ax, Abantu, Ael and Aend.
Weak A type A3 gives a characteristic mixed field agglutination
Group Anti-A Anti-B Anti-A,B Anti-H A cells B cells O cells
pattern when tested against polyclonal anti-A and anti-A,B
antiserum. However, stronger agglutination is observed when
O 0 0 0 4 4 4 0
using most monoclonal blood grouping reagents. Table 6·8
Oh 0 0 0 0 4 4 4
compares reactions between group A and subtypes. Anti-A1

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 73

Table 6·8 Comparison of reactions: group A and subtypes

Reaction of serum/plasma with


Monoclonal reagents reagent red cells

Red cell phenotype Anti-A Anti-B Anti-A,B Lectin anti-A1 A1 A2 B O

A1 4 0 4 4 0 0 4 0
A2 4 0 4 0 + or 0 0 4 0
A3 1 mf 0 1 mf 0 + or 0 0 4 0
Ax Micro+/1 0 1 0 + or 0 0 4 0

may or may not be produced, although it is often produced may contain A-, B- or H-like substances. Additional
by group Ax individuals. Note that type A3 shows mixed field exposure to the antigen can result in more potent antibody
agglutination with anti-A and anti-A,B and that type Ax formation.
reacts macroscopically with monoclonal anti-A,B. This immune response may be prompted by:
• Presence of ABO incompatible fetal red cells in the maternal
circulation during pregnancy and at delivery
Weak AB
• Injection of A or B substances that may be found in vaccines,
In weak AB types, the B antigen may occur with any of the either in the culture medium or in the micro-organisms
weak A subtypes. themselves
• The accidental transfusion or injection of ABO incompatible
red cells.
Weak B
Alloagglutinins that are weak or missing in adults may
Subgroups of group B are suspected when the expression of occur in weak subgroups of A or B, agammaglobulinaemia
the B antigen is weak or cannot be easily detected. Subgroups (patients with low levels of serum globulins), twin chimerism,
of B are rare and are found mainly in populations where the old age or treatment with immunosuppressive drugs.
frequency of group B is high as in African and Chinese
populations. The subgroup cannot be detected if inherited
Alloagglutinins in infants
with a normal B allele. The weak B subgroup may be inherited
with an A allele giving rise to a normal A, weak B phenotype, Alloagglutinins are not normally detected in newborn infants
ABweak. and develop after 3–6 months of life due to exposure to
A- and B-like substances/antigens in the environment. If
ABO antibodies are detected in neonatal blood samples, they
Acquired B
are usually agglutinating IgG antibodies of maternal origin.
This is caused by the action of enzymes which break down Table 6·9 shows the grouping results of a group B newborn
the group A antigen N-acetyl-D-galactosamine to galactos- and an infant of 6 months of age.
amine which is similar to the structure of the group B antigen
immunodominant sugar (D-galactose). Some anti-B reagents
Anti-A1
react with this acquired group B antigen and a group A indi-
vidual could be incorrectly grouped as group AB. It is impor- Individuals of phenotypes A2, A2B and weaker subgroups of
tant to select anti-B grouping reagents carefully to ensure A may have anti-A1 in their serum/plasma. This antibody will
that they do not react with acquired B cells. The condition is react with group A1 cells. Anti-A1 is usually a cold reacting
rare but may be associated with gastrointestinal bacterial antibody, which is not of clinical significance. As it seldom
disease or caused by bacterial contamination of a blood sample. reacts above +25°C, it is unlikely to cause transfusion reac-
The individual’s red cells often become polyagglutinable. tions or HDFN. It may, however, mask a clinically significant
antibody.
Anti-A1 occurs naturally in the serum/plasma of about 2%
ABO system antibodies of A2 individuals and 26% of A2B individuals. The antibody
Healthy adults who lack a particular ABO group antigen occurs more frequently as the strength of the A antigen
on their red cells usually have the corresponding antibody decreases, therefore weak A (or weak AB) individuals are
in their serum as a result of stimulation from the environ- more likely to have anti-A1 in their serum/plasma than A2
ment, such as exposure to certain bacteria or food that (or A2B) individuals.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
74 Blood group systems

Table 6·9 Newborn and infant ABO grouping results

Forward grouping Reverse grouping Interpretation

Age of infant Anti-A Anti-B Anti-A,B A cells B cells O cells Group

Newborn 0 4 4 0 0 0 B
Infant of 6 months 0 4 4 3 0 0 B

• Their naturally occurring anti-A,B antibodies are not


Anti-H (other than Bombay anti-H, -A, -B)
harmful to the red cells of the recipient if whole blood is
As individuals of group A1, A1B and B have very little H antigen transfused, provided the alloagglutinins are ‘low titre’
on their red cells, they sometimes develop anti-H in their (lack ABO haemolysins).
serum. This antibody can be recognized by its strong reaction However, blood from ‘high titre’ group O donors, which
with group O red cells, a weaker reaction with group A2 cells contains immune anti-A and/or -B, may only be transfused
and usually a failure to react with group A1 or group B red into group O recipients (homologous group transfusion). This
cells. Anti-H of this nature, which is formed by individuals is because these ‘dangerous’ universal donors have potent
who are not H deficient, is usually benign and occurs naturally alloagglutinins with haemolysing characteristics, which
in the serum of some group A1, B and A1B individuals. may cause severe haemolytic reactions when infused into
recipients with A, B or AB antigens on their red cells. The risk
of transfusing harmful anti-A and anti-B in blood group O
Group O serum
whole blood can be reduced by the transfusion of group O
Group O serum is not a simple mixture of anti-A and anti-B. red cell concentrates.
It cannot be separated by selective adsorption using either In practice, however, it is better to transfuse a patient with
group A or group B cells and is a cross-reacting antibody blood of the same ABO group (ABO identical) and to conserve
generally known as anti-A,B. Various theories have been stocks of group O blood for group O patients and for
suggested to explain this cross-reactivity (including Wiener’s emergency use.
C theory) and it appears that the anti-A,B produced by group
O individuals detects a structure common to both A and B Universal recipient
antigens. Broadly speaking, group A1B individuals are universal
recipients because:
• They have both A1 (the full quota of A antigen) and B
Clinical significance of the ABO system
antigens on their red cells.
• They usually lack ABO antibodies in their serum.
Clinical significance in transfusion
• Therefore, irrespective of the ABO blood group antigens
Of all the blood group systems, the ABO is the most important on the donor cells, there are no antibodies in the group AB
in transfusion because the alloagglutinins are normally recipient to react with them.
present in the absence of the corresponding antigen. Strong Likewise, provided that the alloagglutinins in the
reactions take place when incompatible bloods are mixed plasma of the donor are low titre, they will not be harmful
with each other, not only in vitro but also in vivo. Even an to the A and B antigens on the red cells of the group AB
initial transfusion of group A blood into a group B patient recipient.
may be disastrous, because the naturally occurring anti-A in
the blood of the group B patient would react promptly
Clinical significance in haemolytic disease of the
with the incoming group A cells, causing agglutination and
fetus and newborn
haemolysis of the donor cells and a likely haemolytic
transfusion reaction. Some individuals produce potent, high titre anti-A and/or
anti-B, consisting of a mixture of IgM and IgG antibodies,
Universal blood donor with haemolysing characteristics in the presence of comple-
Individuals of blood group O are termed universal blood donors, ment. This immune anti-A and/or -B in pregnant women can
as their blood can usually be safely infused into recipients cause ABO HDFN with varying degrees of severity, although
of other ABO groups (heterologous group transfusion) because: the fetus is rarely affected in utero. ABO HDFN typically
• They do not have A or B antigens on their red cells to react develops within a few days of birth. See Section 7: Haemolytic
with antibodies within the circulation of the recipient. diseases, for more information.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 75

Lectins (plant agglutinins) Unique features of ABO system


Certain plant extracts (usually seeds) agglutinate human and The critical unique feature of the ABO blood group system is
animal red cells. Two names have been suggested for these that unlike other blood group systems, the anti-A and/or anti-
plant agglutinins: phytagglutinins and lectins, the latter term B alloagglutinins are invariably present in the serum/plasma of
used for those which show red cell specificity. Note that these every healthy adult when the corresponding antigen is absent.
substances are not antibodies. Some lectins are described in As the ABO antigens are widely distributed throughout the
succeeding discussions: body, the ABO group must be considered in organ transplant.
Some organs, e.g. the kidney, must be ABO compatible. In bone
marrow transplantation, ABO incompatibility is acceptable
Lectin anti-A1
because of the lack of expression of ABO on stem cells,
The most useful lectin anti-A1 is found in Dolichos biflorus: but precautions need to be taken such as removal of the
the extract strongly agglutinates A1 and A1B cells; it reacts unwanted donor red cells or plasma. Note: Anomalous red
less strongly with A2 cells and very weakly with A2B cells. cell typing may be seen post transplantation.
The extract can therefore be standardized by dilution as a
specific anti-A1 reagent.
Practical application
The cornerstone of safe blood transfusion practice is to
Lectin anti-H
transfuse safe blood of the compatible ABO group. It is
Lectin anti-H can be extracted from the seeds of Ulex euro- critical that the ABO group on all samples, whether from a
paeus or the common European gorse. Ulex is invaluable patient or a donor, is correct, as ABO group mistyping can
for the classification of group O secretor/non-secretor saliva have fatal consequences.
(or group O secretor status).

Rh blood group system


ABH secretion The discovery of the Rh groups by Landsteiner and Wiener in
In addition to being present on the red cells, A, B and H 1940, together with the work of Levine and Stetson in 1939,
antigens are present on most other body cells as glycolipids. heralded the greatest discovery in the blood grouping field
Blood group substances of the same ABO group as the red since Landsteiner described the ABO system in 1900.
cells may also be found in the serum/plasma and are readily In 1939 Levine and Stetson described how the mother of a
detectable in the saliva and other body fluids of most stillborn fetus suffered a severe haemolytic reaction when
individuals. transfused with her husband’s blood. The mother, who
The secretor status is controlled by the Se and se genes at obviously lacked some ‘new’ antigen, must have been
the secretor locus. Se is the dominant gene and is responsible immunized by her fetus that possessed this antigen, having
for the secretion of H. Approximately 75% of the general inherited it from the father. When the ABO compatible
population secrete ABH substances (in the form of water- husband’s blood was transfused, the maternal antibody
soluble antigens) in abundance in all their body fluids. The reacted with this same antigen on his red cells.
ABO group of a secretor may be determined by testing the In 1940 Landsteiner and Wiener, having immunized rabbits
saliva to determine the presence or absence of A, B and H with the blood of a rhesus monkey (Macaques mulatta),
substance. The remaining 25% of the population are termed discovered that the resulting antibodies agglutinated not
non-secretors. Table 6·10 shows the soluble antigens secreted only the monkey red cells but also the red cells of about 85%
according to ABO group. of white people.
Later work, however, showed that the red cell antigens
detected by the human-derived antibody and the animal
Table 6·10 soluble ABH antigens according to ABO group antibody were not identical and belonged to two different
blood group systems. The blood group system detected by the
Group Soluble antigens present human-derived antibodies is now known as Rh (not Rhesus)
and the antigen is called D. The antigen originally described
A secretor A and H by Landsteiner and Stetson is LW in the LW blood group
B secretor B and H
system. The two systems are serologically, biochemically and
O secretor Abundant H
genetically different from one another. The locus for the Rh
AB secretor A, B and a little H
Non-secretor Not readily detectable
genes is on chromosome 1 and is linked to the gene for
elliptocytosis. The locus for LW is on chromosome 19.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
76 Blood group systems

It was soon realized that the Rh antibodies produced in


Molecular studies
humans were not as simple as they had first appeared, and that
many sera contained antibodies of more than one specificity. Following studies on the Rh blood group system at the DNA
Many related antigens were found by workers in England level, it has been shown that there are two Rh genes, RHD
and in the United States of America. This led to the discovery and RHCE, at the Rh locus. The RHD gene produces the D
of the five major Rh antigens, D, C, E, c, and e. The Rh blood antigen. At the RHCE gene locus, depending on the allele
group system has now been shown to be one of the most present, one of four alternative antigenic combinations are
complex multi-allelic blood group systems with more than produced, namely ce, Ce, cE or CE. D+ individuals inherit two
50 antigenic specificities having been described. Rh genes: RHD coupled with one of the alleles of RHCE from
each parent. In most D– individuals, RHD is deleted, and
individuals possess the RHCE genes only. As a result, most
Rh genetics and inheritance D– individuals lack the total RhD protein on their red cell
Most individuals are either D+ or D– and the expression or membrane but this does not appear to have an adverse effect
absence of the D antigen on the red cells results from the on the cell function. Note that many apparent D– individuals
presence or absence of an RHD gene. A D+ individual may of African origin have a D antigen, which is not expressed
inherit two RHD genes, one from each parent (homozygous) because of the presence of the RHD pseudogene or inactive gene.
or one RHD gene from either parent (hemizygous). The two The RHD and RHCE locus genes produce separate proteins
pairs of antithetical antigens C and c, and E and e are but are located in the red cell membrane next to each other,
controlled by the various RHCE genes. The RHD and RHCE forming a complex of antigens. As the two gene loci are in
alleles are inherited as a gene complex or haplotype. such close proximity, many of the unusual Rh variants are
Two different theories were initially proposed for the genetics the result of various genetic occurrences within the two gene
and inheritance of the Rh blood group system but these loci, such as unequal crossing over, or mutations. Both the
have been disproved by molecular genetic studies. RhD protein and the RhCcEe proteins comprise 400 amino
acids each. Whereas D– individuals generally lack the entire
RhD protein, the difference between ce and Ce proteins are
Fisher Race theory (UK – theory of three pairs of
differences in four amino acids, and ce and cE proteins differ
linked genes)
by one amino acid.
In 1943 the statistician Fisher, studying the results of Race The presence or absence of the RHD gene, together with
and co-workers in England, noticed that some reactions were one of the four possible alleles of the RHCE gene, results in
antithetical (opposite), and he therefore supposed that eight possible gene combinations or complexes. The Rh
there were three sets of alleles involved: C and c, D and d, E genotype is therefore a combination of any of the eight
and e. Fisher assumed that the three genes, if separable, must be possible haplotypes. One Rh gene complex or haplotype is
very closely linked, for no crossing over had been observed. inherited from each parent.
The CDE nomenclature was devised and although it did not
accommodate subsequent complexities in the Rh system it
Terminology
was easy to use.
Although this theory suggested that antibodies to all the The eight possible gene complexes or haplotypes are shown
antigens described are able to be stimulated in individuals in Table 6·11. The symbol d denotes the absence of D. There
lacking the corresponding antigen, no anti-d has ever been
Table 6·11 Rh genes together with gene complex (haplotype) and
found.
shorthand nomenclature

Wiener theory (USA – theory of multiple Rh genes present


Gene Shorthand
allelomorphic genes)
RHD gene RHCE gene complex/haplotype nomenclature
This is a theory of multiple allelic genes occurring at a single
chromosomal locus (rather than at three closely linked loci). D Ce DCe R1
One gene complex is inherited from the mother and one from D cE DcE R2
the father. It was thought that each gene complex produced D ce Dce Ro
D CE DCE Rz
an agglutinogen which had several serologic specificities (blood
d Ce dCe rl
factors or antigens). One agglutinogen could react with various
d cE dcE rll
antibodies because it had as part of its structure more than
d ce dce r
one antigen. The Rh-Hr nomenclature was developed to d CE dCE ry
describe the gene complexes, agglutinogens and antigens.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 77

is no d gene or d gene product and d therefore represents an RHD Table 6·12 Frequencies of some Rh genotypes in a UK population
deletion or an inactive RHD gene. The eight Rh gene com-
plexes or haplotypes have each been allocated a shorthand Phenotype results using Rh antisera
Rh Percentage
notation, the symbol R indicating the presence of an RHD gene genotype Anti-D Anti-C Anti-E Anti-c Anti-e frequency
and D antigen and r the absence of the RHD gene antigen.
Table 6·11, shows the Rh genes, complexes/haplotypes and R1R1 + + 0 0 + 18
shorthand nomenclature. R2R2 + 0 + + 0 3
R1R2 + + + + + 13
Numerical terminology R1r + + 0 + + 35
In 1962 Rosenfield introduced a new terminology for the Rh R2r + 0 + + + 12
R° + 0 0 + + 2
system based on a numerical system. Each antigen was
rr 0 0 0 + + 15
numbered, as was the antibody detecting it. For example, the
r lr 0 + 0 + + 0·4
D antigen is Rh:1 and anti-D is anti-Rh1. Blood lacking the
r llr 0 0 + + + 0·8
D antigen is noted as Rh:−1 (minus one). This numerical r lr l 0 + 0 0 + < 0·1
terminology lends itself to computerization. It is now the r llr ll 0 0 + + 0 < 0·1
basis for the ISBT terminology for all the blood groups. r lr ll 0 + + + + < 0·1
R zr + + + + + Very rare
Terminology hints r yr 0 + + + + Very rare
The Rh terminology can be very confusing and there are several
different ways of documenting Rh. Below are several points
that may be of assistance when using the shorthand notation:
• Whenever D is present, use the letter R together with the D– based on D typing results which detect the presence or
appropriate number or symbol (see second, third and absence of the D antigen. The frequency of the D antigen
fourth bullet points). varies in different populations, e.g. in a white population
• The C antigen is associated with either 1 or l (prime). If C 85% are D+ and 15% D– but in the Far East it is extremely
occurs with D use 1. When D is absent use l (e.g. DCe = R1 rare to type D–. Ideally a patient or donor sample should be
and dCe = r l). tested with two different anti-D reagents and if the test
• The E antigen is associated with either 2 or ll (double results concur then the sample can be designated D+ or D–.
prime). If E occurs with D use 2. When D is absent use ll As a result of the introduction of potent monoclonal anti-D
(e.g. DcE = R2 and dcE = r ll). typing reagents as routine blood grouping reagents, many
• When C and E are absent but D is present, the notation Ro red cells previously typed as weak D are now typed D+ and
is used. the typical former weak D types cannot be differentiated from
• The phenotypes dCE and DCE are both very rare, so it D+ in routine tests.
seems logical to use y and z to describe them (ry and Rz)! Donor red cells should be typed for weak D either using a
• Genotypes are written in italics, and subscripts become sensitive technique such as the indirect antiglobulin test (IAT)
superscripts (e.g. R2 becomes R2). or by using the appropriate monoclonal blood grouping
reagents, which are known to detect weak D.
Patients should be tested with the appropriate anti-D
Frequencies
reagents. It is not necessary to detect the very weak D
The frequency of the eight possible Rh gene haplotypes varies variants because if patients are typed D–, they should receive
between different populations. For example, the gene com- D– blood (see Rh antigens later in this section).
bination R° is seen more frequently in black people particularly
in sub-Saharan Africa than white people, whereas the Terminology notes
haplotype r is more frequent in white people than in black It should be noted that there are various methods to describe
people. Table 6·12 gives the frequencies of most of the Rh the results of D typing and the Rh type, as indicated in the
genotypes in a UK population. list below:
Result: D+ and D–
Description: D+ and D–
Rh typing RhD-positive and RhD-negative
RhD positive and RhD negative
D typing
Rh-positive and Rh-negative
The D antigen is the most clinically significant blood group Rh positive and Rh negative
antigen in the Rh system. Individuals are divided into D+ or D-positive and D-negative

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
78 Blood group systems

Table 6·13 Variation in frequency of Rh phenotypes: a study of southern African populations

Rh reagent antisera Ethnicity: percentage frequency

Anti-D Anti-C Anti-E Anti-c Anti-e Phenotype White Black Indian

+ + 0 + + DCe/dce R1r 33 13 30
+ + 0 0 + DCe/DCe R1R1 18 <1 48
+ 0 + + + DcE/dce R2r 13 15 4
+ 0 + + 0 DcE/DcE R2R2 2 <1 <1
0 0 0 + + dce/dce rr 14 2 3
+ 0 0 + + Dce/dce Ror 3 66 2

Table 6·14 Rh phenotypes and possible genotype using UK-derived statistics

Possible options Known frequency


Rh reagent antisera
for Rh genotypes (%) of genotype
Anti-D Anti-C Anti-E Anti-c Anti-e Phenotype based on phenotype (white)

+ + 0 + + DCe/dce DCe/dce R1r 33a


DCe/Dce R1Ro 2
Dce/dCe Rorl < 0·1

a
This is the probable or most likely phenotype for white people.

In this publication the terminology D+ and D– is used if the red cell sample is homozygous or heterozygous for the
together with Rh-positive and Rh-negative where appropriate. RHD gene as there is no d gene product. Accurate genotype
The term Rh-positive indicates that the test result is D+. The determination can only be established by the use of molecular
term Rh-negative indicates that the test result is D–. genetic techniques or by informative family studies.
Table 6·14 shows an example of an Rh phenotype and the
possible genotype options, using figures derived from UK
Rh phenotyping
statistics for a white population. The probable genotype can
The Rh phenotype can be determined by typing the red cells then be determined.
with specific reagents; anti-D, anti-C, anti-E, anti-c and anti-e It should also be noted that the presence of the very rare
in the laboratory. Positive and negative test results using variant genes such as --- (Rhnull), Dc– and D– - result in all
the above reagents denote the presence or absence of the Rh or some of the Rh antigens missing from the red cells and this
antigens and this is known as the Rh phenotype. will affect the possible genotype calculations.
Table 6·13 shows the variation in percentage frequency of
various Rh phenotypes in one study of southern African
populations. The symbol d is used to denote the absence of
Rh antigens
the D antigen. The Rh antigens are coded for by the RHD and RHCE genes,
It is not possible to determine the genotype of an individual each of which produces a separate protein that is inserted into
from the red cell phenotype result but the most probable the red cell membrane. The RhD protein crosses the red cell
genotype can be deduced using the haplotype frequency membrane 12 times, giving rise to six extracellular domains.
data. The phenotypes shown in Table 6·13 do not reflect one Despite many studies, the exact function of the Rh proteins
specific genotype. For example, cells which are phenotypically within the red cell membrane is unknown, but their structure
Ror may be genotypically R°R° or R°r. suggests a transmembrane transporter function. The functions
However, as the genotype incidence varies between different of the RhD and RhCcEe proteins appear similar. In cases of
populations and ethnic groups, the ‘probable genotype’ result the very rare type Rhnull, the absence of the Rh proteins has
should be treated with reservation. It is an advantage to know shown that the red cells are abnormal morphologically and
the ethnicity of the individual being typed so that the appro- individuals often suffer from some degree of haemolytic
priate frequencies can be used. The phenotype cannot determine anaemia as the red cells are abnormal in shape.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 79

and RhD classification to describe the Rho mosaic antigen, but


D antigen
this terminology is now obsolete.
The vast majority of all populations are D+. The D antigen is Tippett and Sanger used the Category Classification to
extremely immunogenic and is likely to stimulate antibody describe partial D. Their original classification, using human
production in D– individuals exposed to the D antigen. It is sera, divided partial D bloods into six categories: I, II, III, IV,
the most clinically significant of the Rh antigens and plays V and VI, with category VI containing fewer D epitopes than
a significant role in HDFN (refer to Section 7: Haemolytic the others. The categories were shown to be inherited. The
diseases). original framework of this study has allowed for the addition
of further partial D type complexities, as more information
became available. The use of epitope-specific monoclonal
Weak D (previously termed Du)
anti-D has enabled the partial D to be categorized further
Weak D describes a weaker form of D+, where fewer D antigen into categories II, IIIa, IIIb, IIIc, IVa, IVb, Va, VI, VII and many
sites are present on the red cell as compared with a normal others. Panels of monoclonal anti-D antibodies are now
D+. Studies have shown that D+ red cells of the R1r phenotype available to classify partial D types. The molecular structures
have about 10 000 antigen sites per cell, whereas R2R2 of the many partial D types have been extensively studied
phenotypes have about 30 000 sites. Weak D cells have far and it has been shown that various RHD and RHD-CE-D, and
less than this number, although the number is variable. The RHCE-D-CE hybrid genes give rise to different partial D
weak D characteristic is usually the result of the inheritance types.
of a genetic variation.
The term Du for weak D is now obsolete. It was used to
Clinical significance of weak D and partial D
describe those forms of the D antigen that reacted weakly in
laboratory tests when tested with different polyclonal anti-D Weak D individuals do not usually produce anti-D although
reagents. With the use of monoclonal anti-D reagents many isolated cases have been reported whereas partial D type
weak D types now type as D+ and cannot be distinguished individuals may develop clinically significant anti-D.
from normal strength D+ in a routine laboratory. Patients:
The identification of a weak D type will depend on the • Weak D patients very rarely form anti-D, and it is not
anti-D reagents selected for use and the technique used. considered necessary for weak D mothers who carry D+
If two potent, type IgM agglutinating monoclonal anti-D infants to be given anti-D immunoglobulin.
reagents are used routinely for patient testing, most weak D • As routine anti-D typing reagents do not differentiate
samples will type D+. Only the weakest form of weak D will most partial D type bloods from normal D+ bloods, these
be identified as weak or may be typed as D– and such patients rare individuals may be stimulated to produce anti-D if
will receive D– blood. The weak D type should, however, be transfused with D+ blood. The anti-D can cause severe
detected in donor samples by either proceeding to the second HDFN.
(IAT) phase of a monoclonal blend reagent or by using • A partial D mother will not be identified as a candidate for
agglutinating monoclonal anti-D reagents specifically selected anti-D immunoglobulin. Should she deliver an infant with
for the ability to detect weak D. normal D and have a transplacental haemorrhage, she may
be stimulated to develop anti-D to the missing epitope(s).
Future pregnancies may be complicated by HDFN.
Partial D
Donors:
In 1953 there was a report of a D+ individual who had anti-D • Blood donors should be tested for weak D, and their blood
in the serum/plasma. Since then, many examples of D+ with transfused into D+ recipients.
anti-D have been reported, although overall it is a rare • Most partial D blood donors will be typed as D+.
occurrence.
The term partial D is used to describe the phenotype of
Trans effect of C
those rare individuals, whose red cells lack one or more of the
D epitopes. The D antigen is considered to be a mosaic of A weak D phenotype can occur as a result of the trans effect
epitopes. If some D epitopes are missing, then the individual of C. If the haplotype encoding the D antigen is in trans (on
can make an antibody specific for the missing epitope/s if the opposite chromosome) with a haplotype encoding C but
they are exposed to normal D+ cells. The anti-D produced in not D (e.g. dCe), the expression of the D antigen may be
this way reacts with all normal D+ cells, which have all the weak. Family studies have shown that when the haplotype
epitopes, but fails to react with their own cells and cells of encoding the weak D is not in trans with C, then the D antigen
the same or similar partial D types. is expressed normally. For example, Dce/dCe will type as
In their studies, Wiener and Unger used the RhA, RhB, RhC weak D, whereas DCe/dce may type as normal D.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
80 Blood group systems

blood shortage. The reasoning is that should anti-D be produced,


Cc and Ee antigens
it will not be able to cause HDFN.
The Cc and Ee antigens are less immunogenic than the D Good laboratory practices should ensure that both patient
antigen and may demonstrate dosage effect depending on and donor are correctly Rh typed and that incompatible
the reagent used. A number of variants, particularly variants transfusions resulting from the presence of Rh antibodies are
of the e antigen, have been described such as the rare hrS- avoided.
and hrB- found mainly in black people. There is a risk that patients who lack the C, E, c or e antigens
will be exposed to these antigens during transfusion. This
occasionally leads to the production of the corresponding
Cw antigen and anti-Cw
antibodies, but these antigens are far less immunogenic
Although the Cw antigen was originally thought to be an than the D antigen. For example, a recipient who has the Rh
allele of C, it has been shown to be an allele of the high phenotype DCe/DCe may receive blood from a donor who is
frequency Rh antigen MAR. Cw+ cells are usually C+. Anti- DcE/DcE. In this example, it is possible that the recipient may
Cw is not necessarily produced in response to a known red be stimulated to produce of anti-c and/or anti-E antibodies.
cell stimulus and may occur in combination with other Further transfusions of blood positive for c and/or E antigens
antibodies to low frequency antigens. may cause a haemolytic reaction if the antibody is not
detected during crossmatching.
In the case of multi-transfused patients or patients on long
G antigen and anti-G
term transfusion therapy, Rh phenotyped, matched blood
Red cells that are C+ or D+ are generally G+, although very should be provided i.e. the units selected should be matched
rare exceptions have been reported. The RhC and RhD to the patients’ Rh phenotype. The blood should also be
proteins share an amino acid sequence that is recognized by matched for the K antigen in the Kell blood group system,
anti-G. Many anti-C+D sera may therefore contain anti-G. which is strongly immunogenic. This will prevent antibodies
Anti-G will appear to be Anti-C+D in routine laboratory tests being formed to these antigens, which could complicate
but anti-G can be separated from anti-C and anti-D by future transfusions or pregnancies.
adsorption and elution studies, or identified by the use of When Rh antibodies are present, antigen negative
rare cells. crossmatch compatible blood should be transfused.

Other Rh antigens Clinical significance in haemolytic disease of the


fetus and newborn
The Rh blood group system is a very complex system, and 49
different antigens, including the ones discussed previously, Rh antibodies, particularly anti-D and anti-c, are capable of
are recognized by the ISBT Committee on Terminology for causing HDFN. (See Section 7: Haemolytic diseases for details.)
Red Cell Surface Antigens. Anti-D is the most common cause of severe HDFN and may
be combined with other Rh antibodies, e.g. anti-D+ anti-C.
Although ‘enzyme only’ reacting Rh antibodies may be
Clinical significance of the Rh system
detected (they often have anti-E specificity), the Rh antibodies
Rh antibodies are nearly always immune type IgG antibodies that cause HDFN are type IgG and are best detected using IAT.
that have been stimulated by exposure to foreign red blood A fetomaternal haemorrhage (FMH), either during pregnancy
cells either through pregnancy or transfusion. or during delivery, acts in exactly the same way as a trans-
fusion, although the volume of cells is much smaller. The
mother may be exposed to antigens she lacks, but which the
Clinical significance in transfusion
fetus has inherited from its father. The best example of this is
The antibodies in the Rh blood group system can cause severe the D– mother who has an FMH of D+ blood and develops
transfusion reactions and are second only to the ABO system anti-D, which may cause HDFN in subsequent pregnancies if
in this regard. The transfusion of D+ blood into D– patients the fetus is D+.
should be avoided, as the D antigen is highly immunogenic HDFN usually increases in severity with each succeeding
and can stimulate antibody production in the recipient. D+ pregnancy. The anti-D antibodies increase in titre and
Group O (low titre) is considered to be the universal donor. avidity with every additional immunization of fetal cells, so
The universal donor should also be D– if the blood is to be that eventually they may be of sufficient potency to cause
transfused into D– patients. However, in order to conserve intrauterine death of the fetus. Before the advent of prophylaxis
supplies of D– blood, one may consider transfusing cross- (preventative treatment) with Rh immunoglobulin, anti-D was
matched D+ blood into male D– recipients in cases of severe responsible for about 90% of severe haemolytic disease cases.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 81

Other Rh antibodies, particularly anti-c, may cause severe Table 6·15 MNS blood group frequencies in percentage: excluding U which
HDFN. For example, it is possible for an Rh-positive (DCe/ is a high frequency antigen
DCe) mother who has been stimulated to produce anti-c, to
give birth to an Rh-positive (DCe/dce) infant suffering from Phenotype White Black
HDFN caused by anti-c.
M+N– 28 26
M+N+ 50 44
Unique features of the Rh system M–N+ 22 30
S+s– 11 3
• The Rh blood group system is an extremely complex S+s+ 44 28
blood group system. S–s+ 45 69
• Many variant antigens have been described.
• The antibodies are generally type IgG and are clinically
significant.
• Anti-D is the most common serious cause of severe HDFN.
Although the MN types and Ss types are shown separately
in the table, the two sets of alleles form a single blood group
Practical application
system. The MN antigens are situated on glycophorin A
• It is essential that the laboratory system enables both (GPA) and the Ss antigens on glycophorin B (GPB). There
patients and donors to be correctly Rh typed using the are two amino acid differences between the M and N on
appropriate reagents. glycophorin A and a single amino acid difference between S
• Adequate antibody screening tests must be available to and s on glycophorin B. Amino acid substitutions and hybrid
detect the clinically significant antibodies in patients requir- GPA/GPB glycophorins result in many of the large number
ing transfusions, pregnant women and donor plasma. of antigens identified within this system, e.g. Mg, Miltenberger,
• Anti-D immunoglobulin should be given to all D– women Sta, Dantu and others. The very rare type Mk is the result of
who have not developed anti-D, antenatally and/or post- the absence of both red cell GPA and GPB.
natally, the regime followed depending on the guidelines The S and s antigens are associated with the high frequency
for the particular service/country. antigen, U. The rare phenotype S-s-U− occurs in black
• If D+ blood was given to a D– patient with child bearing populations and a rare variant form of U gives rise to a
potential, whether in emergency or in error, the administra- S-s-U+ phenotype.
tion of anti-D immunoglobulin to the patient should be
considered, the size of the dose depending on the volume
Antibody characteristics
of blood transfused.
• Enzyme treatment of red cells denatures M, N and S
antigens but not s and U, resulting in anti-M, anti-N and
Other major blood group systems anti-S not usually being demonstrable by enzyme
Twenty nine different blood group systems have been recog- techniques.
nized, including the two most important systems just described. • Some examples of anti-M and anti-S are enhanced by
In addition to the red cell antigens described in the ABO and lowering the pH of the serum.
Rh blood group systems, there are many more that can be • M and N antibodies are usually cold agglutinins reacting
detected on human red cells by specific antibodies and these by saline agglutination techniques. The antibodies seldom
are allocated to blood group systems, collections or series. react above +20°C.
The succeeding pages will describe the other major blood • Occasionally, anti-M and anti-N react by IAT.
group systems: MNS, P, Kell, Duffy, Kidd, Lewis, and Lutheran. • Although anti-S may be naturally occurring, it is usually
Other systems of practical importance in the laboratory such a complement-binding type IgG antibody. Anti-S is often
as I will be described thereafter with some comments on the produced together with antibodies to low frequency
other systems/collections. antigens or ‘private’ antigens.
• Anti-s seldom occurs and may be demonstrable at
temperatures below +20°C or by IAT.
MNS system (discovered in 1927) • Anti-U antibodies are usually demonstrable by IAT, but
Well-known antigens: M, N, S, s, U some may react by enzyme methods.
Number of antigens: 46 (2006). • Monoclonal anti-M, anti-N and anti-S reagents are avail-
Table 6·15 provides the MNS blood group frequencies able for red cell typing and a lectin anti-N (Vicia graminea
excluding U. or Vicia unijuga) is available for N typing.

© 2008 The Authors.


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82 Blood group systems

Table 6·16 Frequency of P1 antigen in percentage


Clinical significance
Transfusion Phenotype White Black
• Anti-M and anti-N are generally considered not to be of
clinical significance and have rarely been the cause of P1+ (or P1) 79 94
P1– (or P2) 21 6
transfusion reactions. IAT crossmatch compatible blood is
usually given.
• Anti-S, -s and -U are considered clinically significant
Antibody characteristics
and may cause moderate to severe transfusion reactions.
Crossmatch compatible and antigen negative blood • Anti-P1 is usually a type IgM antibody. It is commonly
should be selected for transfusion. encountered as a cold agglutinin but occasionally reacts
at +37°C. However, some rare examples may bind
Haemolytic disease of the fetus and newborn complement and react by IAT.
• Anti-M and anti-N have occasionally caused mild HDFN • Anti-P1 that binds complement and reacts by IAT may
but are not considered to be of obstetric significance. cause transfusion reactions.
• Anti-S, anti-s and anti-U may cause moderate to severe, • Other antibodies that are not actually part of the P blood
even fatal HDFN. group system are noted here:
• Anti-PP1Pk (previously called anti-Tja) is a rare, potent
antibody found in the very rare type p or minus/minus
Features
individuals. (P is the only antigen in the Globoside system
• The M and N antigens are within the glycophorin molecules ISBT 028.) The antibody reacts at all temperatures by all
of the red cell membrane. methods and is frequently present as a haemolysin. It
• Anti-M and anti-N antibodies are not usually detected in causes transfusion reactions, and is a potential cause of
tests using enzymes such as papain or bromelin. recurrent abortions. It rarely causes HDFN.
• Anti-P is found in the serum of all Pk individuals and
will haemolyse P1 and P2 cells in the presence of com-
Practical application
plement. Anti-P is also found in cases of paroxysmal
• Anti-M and N are not considered to be clinically significant cold haemoglobinuria (PCH). PCH is a haemolytic
when they react at low temperatures. Examples that react disease which occurs mainly in children following a
at +37°C should be considered to be of possible clinical viral infection. The sera from such patients give a positive
significance. Donath-Landsteiner test (see Glossary for details).
• Anti-S and anti-s can cause severe transfusion reactions
and HDFN.
Clinical significance
• U is a high frequency antigen and if a patient with anti-U
requires blood it will be extremely difficult to find random Transfusion
U- donors. A rare donor registry needs to be consulted. Anti-P1 is not generally considered to be clinically significant
even when reactive at +37°C and it is not usually necessary
to select antigen negative blood. The blood crossmatched is
P blood group (discovered in 1927) considered compatible if negative by IAT.
Well-known antigen: P1
Number of antigens: 1 (2006) Haemolytic disease of the fetus and newborn
P1 is the blood group antigen in the P system (ISBT Anti-P1 has not been reported to cause HDFN.
003) and the terminology P2 indicates the lack of P1 antigen.
The associated antigens, P, Pk and LKE, are not included
Features
in the P blood group system but are mentioned here for
convenience. The P and LKE antigens are controlled by • Anti-P1 is usually not of clinical significance.
genes at a different locus to that for P1 antigen. The P • P1 antigen is weakly expressed at birth.
antigen has been allocated to the Globoside Blood • P1 substance can be found in various flatworms, and hydatid
Group System (028). Pk remains in Collection 209, symbol (tapeworm) cysts in sheep livers. P1 substance from avian
GLOB, until more information is available to allocate it to a sources, e.g. pigeon egg white can be used in inhibition tests.
specific system. LKE is also classified in Collection 209, GLOB. • The frequency and avidity of anti-P1 is increased in P1-
The frequency of the P1 antigen varies in different individuals suffering from helminth infestations (parasitic
populations as shown in Table 6·16. worm, e.g. hookworm).

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 83

Red cells of the very rare McLeod phenotype show weakened


Practical application
expression of the Kell high frequency antigens, together with
• The P1 antigen varies considerably in antigenic strength and the para-Kell antigens and the K antigen if present. The
known strong P1+ cells should be used for antibody detection. degree of depression of the different antigens varies.
• Anti-P and Anti-PP1Pk are antibodies to high frequency Depressed Kell system-antigens have also been observed in
antigens and it is extremely difficult to find compatible individuals of the rare Ge:-2,-3 phenotype in the Gerbich system.
blood for patients requiring blood transfusions.

Antibody characteristics
Kell blood group system (discovered in 1946)
• Anti-K and anti-k are usually type IgG antibodies, reacting
Well-known antigens: K and k, Kpa and Kpb, Jsa and Jsb optimally by IAT and some examples react by enzyme
Number of antigens: 28 (2006). techniques.
The Kell blood group system is extremely complex. Twenty- • Some examples of anti-K will react at temperatures below
four Kell system alleles have been identified at the KEL locus +37°C.
on chromosome 7. The Kell system is also associated with the • Although anti-K is usually produced in response to
Kx and the Gerbich blood group systems, which adds to its stimulus by transfusion or pregnancy, ‘naturally occurring’
complexity. cases have been reported and are possibly due to bacterial
The most commonly encountered antigens in the laboratory infections such as Escherichia coli. This type of anti-K is
are the K and k antigens. After the ABO and Rh antigens, the usually transient.
K antigen is the most immunogenic. • Some examples of anti-K react more weakly in tests using
Table 6·17 shows the Kell blood group frequencies. LISS or polybrene.
• If a patient develops anti-k it will be difficult to provide
Table 6·17 Kell blood group frequencies in percentage compatible blood because of the low frequency of k–
blood. However, anti-k is a relatively rare antibody as few
Phenotype White Black individuals are k–.

K+k– 0·2 < 0·1


K+k+ 8·8 3·5 Clinical significance
K–k+ 91·0 96·5
Transfusion
Anti-K and anti-k are clinically significant and can cause
Many different Kell and para-Kell antigens are produced severe haemolytic transfusion reactions and delayed transfu-
and the frequency of the antigens, other than the high and sion reactions.
the low frequency antigens, varies greatly between populations.
The well-known high frequency antigens are k, Kpb and Jsb. Haemolytic disease of the fetus and newborn
The para-Kell antigens are very high frequency antigens Anti-K antibodies differ from the other blood group system
that are absent from Ko cells, but have not been shown to be antibodies that cause HDFN as the antibodies appear to
controlled by genes at the KEL locus, although they are destroy the precursor red cells, causing severe anaemia,
located on the Kell glycoprotein. Depending on the specifi- and often death of the fetus. High bilirubin levels are not a
city, the antibodies react with the corresponding para-Kell characteristic as the precursor cells are destroyed. Amniocentesis
antigens, but not with Ko cells. therefore does not give an indication of the severity of the
The Ko type is the rare null type in the Kell system. Ko red cells disease.
lack all the Kell and para-Kell antigens. Ko patients who have
become immunized frequently produce anti-Ku, an antibody Practical application
that reacts with all cells except Ko cells. This makes the provi- K− or k− (i.e. antigen negative) blood and crossmatch com-
sion of compatible blood for such patients extremely difficult. patible blood should be provided to patients with anti-K or
The chemical, dithiothreitol (DTT), destroys all the Kell and anti-k antibodies respectively. In patients requiring long
para-Kell antigens, so the detection of Kell antibodies is not term transfusion therapy, units matched for K as well as the
possible if the following are used in laboratory tests: various Rh antigens should be provided.
• DTT to determine whether IgM or IgG antibodies are
present in a sample.
Kpa and Kpb
• A ZZAP solution that contains DTT and an enzyme such
as papain or bromelin (as used for auto adsorptions). See The Kpa and Kpb antigens result from the presence of two
Glossary for information on ZZAP. allelic co-dominant genes in the Kell blood group system. The

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
84 Blood group systems

Kpa antigen is a low incidence antigen occurring in about Table 6·18 Duffy blood group frequencies in percentage
less than 2% of individuals. The Kpb antigen is a high
incidence antigen occurring in more than 98% of individuals. Phenotype White Black
The antibodies are type IgG, reacting optimally by IAT and
Fy(a+b–) 20 9
are clinically significant both in transfusion and HDFN. The
Fy(a+b+) 47 1
provision of Kp(b−) blood to patients who have become
Fy(a–b+) 33 22
immunized is difficult as most donors type Kp(b+). Antigen
Fy(a–b–) 0 68
negative and IAT crossmatch compatible blood should be
transfused when the intended recipient has the corresponding
antibodies. The Fya and Fyb antigens are destroyed by the action of
proteolytic enzymes. The antigens also progressively deteri-
orate within a short period of time when the cell suspensions
Jsa and Jsb
are prepared in some saline or LISS solutions. The antigens
The Jsa antigen is found mainly in individuals of African are, however, well preserved in anticoagulant solutions and
origin. The incidence is approximately 20% in black people preservation solutions for red cells.
and less than 0.1% in white people. The Jsb antigen is a high The Duffy protein spans the red cell membrane nine times
incidence antigen found in more than 98% of individuals. and is 338 amino acids in length. The difference between Fya and
The antibodies are type IgG and react best by antiglobulin Fyb is caused by one amino acid substitution within the protein.
techniques. Anti-Jsa is seldom seen. All examples of anti-Jsb
have been detected in black people. Both antibodies have
Antibody characteristics
caused HDFN and transfusion reactions. Antigen negative
and IAT crossmatch compatible blood should be transfused. • Anti-Fya and anti-Fyb are type IgG and can be stimulated
If the rare type Js(a+b−) blood is required it may be obtained by transfusion and pregnancy.
through a rare donor registry. • Anti-Fya and anti-Fyb antibodies react by antiglobulin
technique and are not detected by enzyme techniques.
• Anti Fya is often detected together with other red cell anti-
General comment
bodies, particularly Rh antibodies.
The antibodies to the Kell system are usually of clinical • Anti-Fyb antibodies are not frequently encountered.
significance and may cause severe or delayed transfusion reac-
tions. The respective antigen negative and IAT crossmatch
Clinical significance
compatible blood should be transfused. If the antibody is to
a high frequency antigen in the Kell blood group system it The Duffy system antibodies are capable of causing severe
may be necessary to obtain the antigen negative blood from transfusion reactions and HDFN. Antigen negative and IAT
a rare donor registry. crossmatch compatible blood should be transfused to
patients with antibodies.

Duffy blood group system (discovered in 1950)


Well-known antigens: Fya, Fyb
Kidd blood group system (discovered in 1951)
Number of antigens: 6 (2006). Well-known antigens: Jka, Jkb
A pair of co-dominant allelic genes located on chromosome Number of antigens: 3 (2006).
1 at the Duffy blood group locus is responsible for the A pair of co-dominant allelic genes located on chromo-
production of the Fya and Fyb antigens. some 18 at the Kidd blood group locus is responsible for the
The frequency of the Duffy blood group system antigens production of the Jka and Jkb antigens.
varies greatly between different populations. The gene Fy The frequency of the Kidd blood group system antigens
which is a silent recessive gene is very rare in white popula- varies between different populations. The Jka antigen occurs
tions but occurs frequently in black African populations, more frequently in black populations.
particularly in malaria areas. A homozygous expression of The rare Jk(a–b–) or Jk:3 phenotype (the null type in the
the Fy gene results in the Fy(a−b−) phenotype and it has been Kidd blood group system), has been reported in some
shown that these red cells are resistant to invasion by the Polynesian, South American and South African Indian
malaria parasites Plasmodium vivax and Plasmodium know- populations. The urea lysis test is a useful test for screening
lesi. It is therefore an advantage for individuals living in for the rare Jk:−3 phenotype as Jk:−3 cells, unlike Jk(a+) and/
malaria areas to lack the Duffy red cell antigens. Table 6·18 or Jk(b+) cells, are resistant to lysis by 2 M urea.
shows the Duffy blood group frequencies. Table 6·19 shows the Kidd blood group frequencies.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 85

Table 6·19 Kidd blood group frequencies (approximate) in percentage Table 6·20 Summary of Lewis, ABH and secretor gene interactions

Phenotype White Black Asian Genes


Phenotype present Comment (ABH) Comment (Lewis)
Jk(a+b–) 26 51 23
Jk(a+b+) 50 40 27 Le(a+b−) Le sese ABH non-secretor Le gene product acts on
Jk(a–b+) 23 8 49 H precursor substance
Jk(a–b–)/Jk: –3 Very rare Very rare <1 Le(a–b+) Le Se ABH secretor Le gene product acts on
secreted H substance
Le(a–b–) lele Se ABH secretor 9
No Lewis genes are present
Le(a–b–) lele sese ABH non-secretor8

Antibody characteristics
• Anti-Jka and anti-Jkb are type IgG antibodies that activate
Table 6·21 Lewis blood group frequencies in percentage
complement and are stimulated by transfusion or pregnancy.
• The antibodies react best by antiglobulin technique, Phenotype White Black
although some weak examples react by enzyme techniques.
Some examples may be complement dependent and require Le(a+b–) 22 23
the presence of complement before they can be detected. Le(a–b+) 72 55
• They are relatively rare antibodies and are most often Le(a–b–) 6 22
present in serum containing other blood group antibodies.
• The avidity and the titre of the antibodies often diminish
rapidly and the antibodies may not be demonstrable in
subsequent samples from the patient. Lewis antigens are not actually red cell antigens but are
• Anti-Jk3 can be a potent type IgG antibody detected by adsorbed onto the red cells from the plasma. The presence or
IAT. absence of the Lea and Leb antigens is determined by genes
at three different loci:
• H (or FUT1) gene: responsible for production of H substance
Clinical significance
(the precursor substance for the A and B antigens).
Transfusion • Se (or FUT2) or secretor gene: enables the A, B and H
• Both anti-Jka and anti-Jkb (including weak examples) antigens to be secreted.
have caused severe haemolytic transfusion reactions and • Le (or FUT3) gene: the symbol le is used to show the
delayed transfusion reactions. Antigen negative and IAT absence of the Le gene.
crossmatch compatible blood should be transfused to When the Le gene is inherited, the enzyme produced reacts
patients with antibodies. with one of two possible H substrates. The preferred substrate
• If further transfusions are required at a later date the is secreted H substance. This is produced when both H and Se
antibodies may be difficult to detect at the time of the genes are present. The secreted H substance is then converted
crossmatch, or may no longer be demonstrable. Antigen into the Leb antigen. If the secreted H substance is not present
negative blood should be transfused to prevent a delayed as a result of the inheritance of the sese genes (or because of
transfusion reaction. Good records of patients with clini- the inheritance of the very rare hh genes or Bombay Oh gene),
cally significant antibodies should be kept so that the the enzyme product of the Le gene acts on the precursor H
appropriate blood can be crossmatched. substance instead, giving rise to the Lea antigen.
• Anti-Jk3 should be considered to be clinically significant. If no Le gene is present, neither the Lea or Leb antigens are
Compatible rare type Jk:−3 blood may need to be obtained produced, resulting in the Le(a−b−) phenotype.
from a rare donor registry. Table 6·20 provides a summary of the Lewis blood groups,
together with their interactions with the ABH and secretor genes.
Haemolytic disease of the fetus and newborn The Lea and Leb antigens are adsorbed onto the red cells
The antibodies rarely cause HDFN. from the plasma, but the antigenic strength is variable and
may change, e.g. pregnant women often type Le(a−b−). Cord
cells type Le(a−b−) as the red cell Lewis antigens only develop
Lewis blood group system (discovered in 1946) during the first 12–15 months after birth.
Well-known antigens: Lea, Leb The Le(a−b−) phenotype occurs more frequently in black
Number of antigens: 6 (2006). people than in white people, as shown in Table 6·21.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
86 Blood group systems

Table 6·22 Lutheran blood group frequencies in percentage


Antibody characteristics
• Anti-Lea and anti-Leb are often type IgM naturally Phenotype White Black
occurring antibodies. They are usually developed by type
Lu(a+b–) <1 <1
Le(a−b−) individuals. Lu(a+b+) 7 7
• The antibodies generally react best at temperatures below Lu(a–b+) 93 93
+37°C and react by saline and enzyme techniques. Some Lu(a–b–) Very rare Very rare
examples of anti-Lea and anti-Leb are type IgG and react
by IAT.
• Strong reacting type IgG antibodies may cause in vitro
haemolysis in the presence of complement. In addition to the two major antigens, Lua (low frequency)
• Some anti-Leb antibodies react better with group O or group and Lub (high frequency), there are three further pairs of allelic
A2 Le(b+) cells than with group A1 or group B Le(b+) cells. co-dominant genes that control the Lu6 and Lu9 antigens,
These are termed anti-LebH as they react with cells with the Lu8 and Lu14 antigens, and the Aua and Aub antigens. Ten
most H antigen. Anti-LebL does not show a variation in reac- other high frequency antigens are part of the system. No
tion based on the ABO group of the Le(b+) cells being tested. Lutheran antigens are demonstrable by routine laboratory
testing on samples of the rare Lu(a−b−) type or null type.
The Lutheran locus is part of a linkage group situated on
Clinical significance
Chromosome 19, which includes the Secretor, Lewis, H and
Transfusion LW loci.
• The majority of Lewis antibodies do not cause transfusion Table 6·22 shows the Lutheran blood group frequencies.
reactions. The expression of Lutheran antigens is variable, and
• Generally patients with anti-Lea and anti-Leb which reacts dosage effects may be seen.
at +37°C can be transfused with crossmatch compatible
blood. There are, however, some rare examples of Lewis
Antibody characteristics
antibodies, which react strongly at +37°C by IAT and
activate complement. These antibodies should be consid- • The antibodies may be stimulated by pregnancy or
ered to be of potential clinical significance. transfusion.
• Anti-Lua is usually a type IgM antibody reacting by saline
Haemolytic disease of the fetus and newborn techniques. Reactions with anti-Lua often show a typical
Lewis antibodies do not cause HDFN as fetal and newborn ‘mixed field’ result.
red cells lack Lea and Leb antigens. • Anti-Lub is usually a type IgG antibody reacting best by IAT.
• Anti-Lua is seldom seen and rarely causes a problem in the
crossmatching laboratory as compatible blood can be
Features
easily found.
The Lewis antigens are adsorbed onto the red cells from the plasma.
Clinical significance
Practical application
• Most examples of Lewis antibodies are not of clinical Transfusion
importance but there are rare exceptions that are clinically • Both antibodies have been reported to cause mild or
significant. delayed transfusion reactions.
• It is not usually necessary to provide antigen negative • IAT crossmatch compatible blood should be transfused.
blood for transfusion. • The provision of Lu(b−) blood may be difficult but the
• Le(a−b−) screening cells are useful when screening the sera antibody is seldom seen.
of pregnant women for clinically significant antibodies,
as Lewis antibodies (which are not of obstetric significance) Haemolytic disease of the fetus and newborn
will then not be detected. Lutheran antibodies have not been reported to cause severe
HDFN as the antigens are only weakly expressed on cord cells.

Lutheran blood group system (discovered in


1945) I blood group system (discovered in 1956)
a b
Well-known antigens: Lu , Lu Antigen: I
Number of antigens: 19 (2006). Number of antigens: 1

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Introduction to Blood Transfusion Technology 87

Although the I blood group system is not one of the major As the frequency of antigens may vary between population
blood group systems it is included in this section as it is of groups it may be informative to know the ethnicity of the
practical importance. The I antigen is found on all normal patient particularly when trying to find compatible blood for
adult red cells. The expression of I antigen varies with age and a patient with antibodies to an apparent high frequency antigen
with disease, and the degree of expression varies considerably or antigens. Such samples are usually referred to a reference
between individuals. laboratory.
All cord cells type I−, i+ as the I antigen is not developed Antibodies to high frequency antigens are usually detected
at birth. Cord cells may then be used as a source of I− cells for when the serum/plasma reacts with all samples tested but the
antibody identification tests. During the first 2 years of life, autoantibody control is negative. This indicates the presence
the I antigen develops and replaces the i antigen, possibly of an alloantibody, not an autoantibody. Further tests are
because of the action of a transferase enzyme using the I required to identify whether the antibodies are directed
antigen as a substrate. against a high frequency antigen or whether they are a
mixture of antibodies of different specificities. Antibodies to
high frequency antigens that do not form part of a blood
Antibody characteristics
group system include anti-Vel, anti-Lan, and anti-Sda.
• Anti-I antibodies are usually type IgM antibodies that Antibodies to the high frequency antigen, Kna, may be
occur frequently as cold reacting autoantibodies or cold referred to as high titre low avidity (HTLA) antibodies. Chido/
agglutinins. They are seldom seen as an alloantibody. Rodgers antibodies to the high frequency antigens Chido/
• However, potent auto-anti-I cold reacting antibodies can Rodgers react with the complement C4 protein, which is
cause cold autoimmune haemolytic anaemia. found on red cells. The identification of these antibodies may
• The antibodies may be detected at +37°C by enzyme or IAT be time consuming and not all are clinically significant.
methods but they are not normally clinically significant. The antibodies may be directed against the high frequency
• Anti-i is occasionally found in patients recovering from antigens mentioned previously in the various blood group
diseases such as infectious mononucleosis (glandular fever). systems. Many of these antibodies are clinically significant
• Anti-I may be associated with anti-H, forming antibodies and can cause severe transfusion reactions and HDFN. It may
with anti-HI specificity. be extremely difficult to find compatible, antigen negative
blood and donations may have to be obtained from a local,
national or international rare donor registry/frozen storage
Clinical significance
facility. Family members who would be suitable donors may
Transfusion also be tested to see if they lack the particular high-frequency
• Although most anti-I antibodies are not clinically signif- antigen, as siblings are more likely to be compatible than
icant, some examples of auto-anti-I with a wide thermal donors in the random population.
range can be seen in cold haemagglutinin disease. Should If the patient has an antibody to a low frequency antigen,
these patients require blood transfusions, they may be the provision of compatible blood should not be a problem.
given the least incompatible blood, warmed in a validated Antibodies to low frequency antigens may be found by
blood warmer before infusion. chance when crossmatching random units of blood or may
• The patient’s samples may be difficult to type if the be detected when using a commercial panel that includes a
patient’s cells are autoagglutinated. The red cells may panel cell sample positive for a low frequency antigen.
need to be washed with warm saline before testing. Table 6·23 lists some of the rare phenotypes associated
• Crossmatching must be very carefully performed to ensure with different ethnic groups and the increased likelihood of
that the auto-anti-I is not masking clinically significant patients producing antibodies of the corresponding specificity.
antibodies. Note that this should be used as a guideline only as there are
always exceptions.
Haemolytic disease of the fetus and newborn
I system antibodies have not been implicated in HDFN.
Polyagglutination
This is defined as the agglutination of red cells by serum/
Additional blood group systems/collections/ plasma from most normal adult humans, irrespective of the
antibodies reacting with high and low frequency ABO groups involved.
antigens Polyagglutination may be inherited or acquired. Acquired
The blood group systems discussed so far in this section are polyagglutination can be caused by the action of microbial
either of clinical or practical importance. However, patients enzymes on the red cells, as a result of infections such as
may produce antibodies to many other blood group antigens. septicaemia, and in bowel, gastric and respiratory infections

© 2008 The Authors.


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88 Blood group systems

Table 6·23 Rare phenotypes with different ethnic groups and increased The HLA antigens are the products of the HLA-A, -B, -C,
likelihood of antibody specificity -DR, -DQ and -DP genes. The original work on HLA was
performed by serological techniques but now HLA loci genes
Increased likelihood of rare type are detected using DNA assays.
Ethnic Blood
group group system Red cell phenotype Antibody specificity The HLA-A, -B, -C antigens are termed the class I antigens
and the HLA-DR, -DQ and -DP antigens, class II antigens,
Black Rh hrS- Anti-hrS based on their biochemical structure.
Rh hrB- Anti-hrB
Some of the HLA antigens share the same epitope and give
MNS U- Anti-U
rise to the many subtypes or splits. As many different
Kell Js(a+b−) Anti-Jsb
terminologies were in use, the World Health Organization
Indian H Bombay Oh Anti-H, -A, -B
IN In(a+b−) Anti-Inb
(WHO) Terminology Committee was established in 1967 to
Kidd Jk(a–b−) Anti-Jk3 review and standardize the nomenclature.
White Kell K+k− Anti-k As the HLA antigens are found on white blood cells and
Kell Kp(a+b−) Anti-Kpb platelets, and also on tissue cells, this complicates the
Vel Vel- Anti-Vel successful grafting of transplanted organs.
Yta Yt(a−b+) Anti-Yta

Disease association
The HLA-A and HLA-B (class I) antigens are expressed on all
nucleated cells in the peripheral blood. The HLA-DR and
among others. This action can result in the cleavage of sialic HLA-DQ (class II antigens) are not expressed on granulocytes,
acid from glycoproteins and glycolipids, thus exposing the unstimulated T-lymphocytes or on platelets. However,
crypt antigens on the red cells that are not normally able to both play a major immunological role in the defence against
be detected. This is known as T-activation. T-activated cells disease.
are agglutinated by most normal sera as most sera contain The association between certain HLA phenotypes and the
IgM antibodies to these crypt antigens. T-activation can be increased occurrence of a specific disease is statistically
readily detected using the lectin Arachis hypogaea or ‘peanut significant, e.g. HLA-B27 and ankylosing spondylitis, and
anti-T’. Acquired polyagglutination is usually transient, there are many other examples.
unlike inherited polyagglutination.
An example of inherited polyagglutination is the agglutina-
Transplantation
tion of red cells of the individuals of the rare Sd(a+++) or Cad
type. The Sda antigen occurs frequently and the strength of When foreign tissue is transplanted into an individual, the
the antigen varies greatly with the strongest expression being HLA antibodies formed as a result of the transplantation may
seen in the rare Cad phenotype red cells. lead to graft rejection in the case of a solid organ transplant,
or graft versus host disease in the case of a bone marrow
transplant. It has been shown that HLA matching between
Human leucocyte antigen system the donor and recipient lessens the degree of rejection and
The human leucocyte antigen (HLA) system, also know as the improves the survival rate. Both donor and recipient
Major Histocompatibility Complex (MHC) is an extremely should be typed and matched for HLA-A, HLA-B, HLA-DR
complex, polymorphic system. and HLA-DQ and depending on the type of transplant,
The HLA antigens are produced as a result of genes located for ABO.
on the short arm of autosome number 6. The genes are As the HLA system is extremely polymorphic the HLA
located at what is known as the MHC locus. The MHC locus types of recipients and/or possible donors are entered onto
also contains the genes for some cytokines and various local or national registers to enable the best match to be
complement components. Gene products are found on the found for a specific patient.
surface membranes of all nucleated cells, including solid
organs, lymphocytes, granulocytes, monocytes and also on Solid organ transplant
platelets although they are not nucleated. HLA antigens are The patients are HLA typed and the results entered in a
not generally detected on red cells (as mature red cells are not national registry. ABO matching is critical due to the presence
nucleated), and red cells are not suitable for HLA typing. of ABO antigens on body tissue. (It is not necessary for bone
However some HLA haemagglutinins, also known as Bg marrow transplantation as ABO groups are not expressed on
antibodies, may detect certain HLA antigens when expressed stem cells.) The HLA types of the patients are matched against
on the red cells. the HLA type of each donor that becomes available. A

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 89

pre-transplant microlymphocytotoxic crossmatch can be The platelet antigens are located on the platelet membrane.
performed to see if the patient has pre-existing antibodies to Serological tests were originally used to study platelet antigens
the donor organ that would preclude transplantation. but it was extremely difficult to obtain specific antisera for
the tests and many antisera also contained HLA antibodies.
Allogeneic bone marrow transplants With the development of molecular testing, it is possible to
Potential donors are HLA typed in advance, and their results determine the HPA genotype and many studies have now
recorded on a donor panel. The donor panel can then be been performed.
searched to find the best HLA match for a potential recipient. The HPA-1a and HPA-5b are considered to be the most
Although there may be many thousands of donors listed on the immunogenic platelet alloantigens in white people and most
donor panel it is extremely difficult to find a matched donor. cases of FMAIT are the result of anti-HPA-1a.
In cases where patients develop platelet refractoriness
during transfusion therapy, it may be necessary to provide
Transfusion
both HLA and HPA matched blood products.
If a patient is transfused with unfiltered blood products
containing white cells and/or platelets, the patient may
become immunized to foreign HLA antigens. If the patient is
Fetomaternal alloimmune thrombocytopaenia
transfused subsequently he/she may suffer febrile reactions. • The most common cause of severe thrombocytopaenia in
For this reason leucocyte-poor products should be transfused the fetus and newborn is FMAIT.
to patients on long term transfusion therapy. In some cases • Although FMAIT has been regarded as the platelet equiv-
it may be necessary to transfuse immunized patients with alent of HDFN, it occurs frequently during first pregnancies.
HLA-matched blood products. The introduction of leucodeple- In FMAIT, the antigen that stimulates the maternal antibody
tion as a routine procedure in many services has reduced the response is found on fetal platelets where the fetus has
exposure to HLA antigens. inherited an HPA type from the father, lacking in the mother.
• The most common antigens to stimulate the production of
platelet antibodies are HPA-1a and HPA-5b which are the
Pregnancy
most immunogenic platelet antigens. FMAIT can also be
HLA antibodies are more frequently seen in women following caused by antibodies to low frequency HPA antigens.
pregnancy. White cell antibodies in multiparous blood donors • Paternally inherited platelet antigens, lacking in the
have been responsible for transfusion related acute lung mother, may stimulate the production of IgG maternal
injury (TRALI) in transfusion recipients. See Section 14: antibodies as a result of FMH. These IgG antibodies are
Benefits and risks of transfusion for more details on TRALI. able to cross the placenta and sensitize fetal platelets,
causing thrombocytopaenia.
• The diagnosis is usually made after birth when the neonate
Parentage testing
shows symptoms of thrombocytopaenia, and platelet
The HLA system is a highly polymorphic and well-defined specific antibodies are detected in the maternal serum.
system. HLA typing is a useful tool in the range of laboratory • Because of severe thrombocytopaenia, fetal/neonatal
tests used in parentage testing. As the frequency of the HLA intracranial haemorrhage may occur.
genes varies greatly between different populations, the • The antenatal administration of intravenous immunoglob-
ethnicity of the individuals concerned must be considered ulin (a non-invasive procedure) to the mother may lead to
when interpreting results. an increase in circulating platelets in the fetus, reducing
the likelihood of fetal intracranial haemorrhage. The reason
for this is perhaps the modulation of maternal antibody
Human platelet antigen systems production that may be induced by the administration of
The molecular basis for the human platelet antigen (HPA) immunoglobulin.
systems has been determined and a total of 24 platelet- • Although the introduction of antenatal screening has
specific alloantigens have been defined serologically. The been considered, there is generally a lack of optimal
systems are very polymorphic and the frequency of the genes antenatal treatment. The safety of fetal blood sampling
varies in different populations according to the ethnic and platelet transfusion is also a concern. Most laboratories
backgrounds. Exposure to foreign platelet antigens following are unable to predict even severe cases of FMAIT.
pregnancy, transfusion, or organ transplant can result in • As in the case of HDFN, premature induction of labour
alloimmunization. This can be responsible for organ transplant may also be recommended. It appears that caesarean
rejection or fetomaternal alloimmune thrombocytopaenia section may be preferred to natural birth, in order to avoid
(FMAIT). cranial trauma at the time of delivery.

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90 Blood group systems

• The practical problem is that most cases are diagnosed • HLA antibodies are more frequently seen in women
postnatally. Time is important as the condition should be following pregnancy. White cell antibodies in multiparous
treated promptly, preferably with HPA compatible platelets blood donors have been responsible for TRALI in transfu-
although random donor platelets, or (rarely) washed and sion recipients.
irradiated maternal platelets, may be used. Most transfusion • HLA typing is a useful tool in the range of laboratory
services do not have HPA matched platelets available as tests used in parentage testing. As the frequency of the
this would require the use of a panel of HPA typed donors HLA genes varies greatly between different populations,
available on call. the ethnicity of the individuals concerned must be
considered.
• The human platelet antigen (HPA) systems are very
Summary of section: Blood group systems polymorphic and the frequency of the different genes
Table 6·24 summarizes the characteristics of the blood varies in different populations.
group systems and their corresponding antibodies. • Exposure to foreign platelet antigens following preg-
Haemolytic transfusion reaction has been abbreviated as nancy, transfusion, or organ transplant can result in
HTR. alloimmunization.
• The HPA-1a and HPA-5b are considered to be the most
immunogenic platelet alloantigens in white people and
Additional points for summary of section: most cases of FMAIT are the result of anti-HPA-1a.
Blood group systems • In cases where patients develop platelet refractoriness
• The human leucocyte antigen (HLA) system, also know as the during transfusion therapy it may be necessary to provide
major histocompatibility complex (MHC) is an extremely both HLA and HPA matched blood products.
complex, polymorphic system. • The most common cause of severe isolated thrombocyto-
• The HLA antigens are produced as a result of genes paenia in the fetus and newborn is FMAIT. Although it
located at the short arm of autosome 6, which is known as has been regarded as the platelet equivalent of HDFN,
the MHC locus. FMAIT occurs frequently during first pregnancies.
• The MHC locus also contains the genes for some cytokines • The diagnosis is usually made after birth when the neonate
and various complement components. shows symptoms of thrombocytopaenia and platelet
• The HLA antigens are the products of the HLA-A, -B, -C, specific antibodies are detected in the maternal serum.
-DR, -DQ and -DP genes. • The practical problem is that most cases are diagnosed
• As the HLA antigens are found on white blood cells and postnatally. Time is important as the condition should be
platelets, and also on tissue cells, this complicates the treated promptly with HPA compatible platelets. Most
successful grafting of transplanted organs. transfusion services do not have HPA matched platelets
• It has been shown that the association between certain available.
HLA phenotypes and the increased occurrence of a specific
disease is statistically significant, e.g. HLA-B27 and
ankylosing spondylitis.
Additional learning activities
• When foreign tissue is transplanted into an individual, (1) It is suggested that students use a medical dictionary
the HLA antibodies formed as a result of transplantation and/or the Internet to clarify the meaning of words and
may lead to graft rejection in the case of a solid organ phrases and to add to the information provided in this
transplant, or graft vs. host disease in the case of a bone section. A list of key words that may be useful in this regard
marrow transplant. is provided below. It is also suggested that the student
• As the HLA system is extremely polymorphic it is difficult compare the published data on the Rh phenotypes and
to find a match for transplantation. genotypes and note the variation in frequencies amongst
• For solid organ transplants the HLA types of recipients are the different populations and ethnic groups.
entered onto local registers to enable the patient to be • ISBT Committee for Red Cell Surface Antigens
matched against a donor as available. • Rh inheritance
• For allogeneic bone marrow transplants volunteer donors • Rh immunoglobulin
are HLA typed and their results entered on a national reg- • HDFN
ister. They are then matched against potential recipients • Rare donor registry
but it is extremely difficult to find a match. • HLA typing
• The introduction of leucodepletion as a routine procedure • Transplantation
in many services has reduced the patient’s exposure to • Class I and class II antigens
HLA antigens. • Graft rejection

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
Introduction to Blood Transfusion Technology 91

Table 6·24 Some of the main characteristics of blood group systems and the corresponding antibodies

System Antibody Predominant type Clinical significance Comments

Anti-A
Can cause fatal HTR and severe HDFN.
Anti-B IgM/IgG HTR and HDFN
ABO Antibodies may be haemolytic
Anti-A,B
Anti-A1 Mainly IgM Rare Found in A2 individuals
Anti-H Mainly IgM Low risk Weakest reactions with group A1 and B cells
H
Anti-H, -A, -B IgM/IgG HTR and HDFN Bombay Oh type
Anti-D Main cause of HDFN
Mainly IgG
Anti-C Often occurs with anti-D
Anti-E IgG/IgM May be naturally occurring and react with enzymes only

Rh HTR and HDFN Severe HDFN


Anti-c
May be produced with anti-E
IgG Not often seen.
Anti-e If individual is a variant e type, antibodies may appear to
be anti-e but individual types e+.
Anti-M Rare HTR Antigen denatured by enzyme treatment
IgM
Ant-N Low risk May show dosage

Anti-S HTR and HDFN rare Antigen denatured by enzyme treatment


MNS IgG/IgM
Anti-s HTR and HDFN Rare antibody
Antibody to high frequency antigen.
Anti-U IgG HTR and HDFN
Compatible blood from rare donor registry
Common cold agglutinin produced by P1− individuals.
P1 antigen Anti-P1 IgM Low risk
P1 substance used in inhibition tests
Anti-Lua IgM mainly Low risk Low frequency antigen
Lutheran
b
Anti-Lu IgG/IgM HTR and HDFN rare Antibody reactions and clinical significance variable
Anti-K Can cause death in utero
Rare antibody
Anti-k
Difficult to find compatible blood
Anti-Kpa Rare antibody
Kell IgG HTR and HDFN
Anti-Kpb Rare antibody. Compatible blood from rare donor registry.
a
Anti-Js Very rare antibody
Antibody to high frequency antigen. Compatible blood
Anti-Jsb
from rare donor registry
Anti-Lea Antigen adsorbed from plasma onto red cells
IgM usually Antigen loss in pregnancy
b
Lewis Anti-Le Type IgG: rarely causes HTR
a b
Anti-Le + Le IgM/IgG Produced by Le(a−b−) individuals
a
Anti-Fy HTR and HDFN Antigen denatured by enzyme treatment
Duffy IgG Antigen denatured by enzyme treatment
Anti-Fyb HTR
Rare antibody
Anti-Jka
Kidd IgG HTR, delayed HTR, and HDFN May require complement for IAT detection
Anti-Jkb

Often detected. Seldom clinically significant. Soluble I


I Anti-I IgM usually Low risk
antigen is secreted in human milk

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92
92 Blood group systems

• HPA typing proceed if the antibody reacts strongly with all the reagent
• Polymorphic panel cells tested? How would knowing the patient’s ethnicity
(2) Use knowledge of the different blood group systems to and/or history assist in the identification of the antibodies
design a set of screening cells for red cell antibody detection and finding compatible blood? Does the service/organization
in the crossmatch laboratory, using donations from the have access to a rare donor registry? If so, what rare blood
blood transfusion service. Create a mini panel as for the types are available? If rare type blood is not available discuss
panel cells decode using the selected screening cells. Which other options to be considered.
combinations of red cell phenotypes are easily obtainable?
Which red cell phenotypes would be difficult to obtain
Conflicts of interest
and what are the reasons for this? What alternatives could
be used? Each contributing author of this section on Blood group
(3) If a strong IAT reacting antibody is detected in the systems (ES and BA) has not received grants, speakers fees
crossmatch laboratory what facilities are available within etc., from any commercial body within the past 2 years. These
the laboratory to identify the antibodies? How should one authors have no potential conflicts.

© 2008 The Authors.


Journal compilation © 2008 Blackwell Publishing Ltd. ISBT Science Series (2008) 3, 68–92