THE ABO BLOOD GROUP
SYSTEM
Chapter 6
KARL LANDSTEINER
• Opened the doors to blood banking
• Took blood from himself and 5 others
• Mixed cells with each other’s serum
and observed reactions.
• First forward and reverse group typing
ABO SIGNIFICANCE
• One of the most important Blood Group Systems
• Causes major transfusion problems with cell incompatibility
• The only blood group system that has antibodies to antigens that
are ABSENT from their RBC’s
ABO TESTING:
FORWARD AND REVERSE GROUPING
• Forward grouping:
• Using known sources of reagent antisera to detect antigens on a
person’s RBC’s
• Anti-A and Anti-B
• Reverse grouping:
• Using known sources of reagent RBC antigens to test for the
person’s ABO group antibodies
• Reagent cells A1 and B
FORWARD TYPING
KNOWN ANTISERA
+
UNKNOWN PATIENT CELLS
REVERSE TYPING
KNOWN RBC’S
+
UNKNOWN PATIENT SERUM
GRADING
REACTIONS: TUBES
4+: One solid button/Clear
background
3+: 2-3 large buttons/Clear
background
2+: Multiple smaller buttons
1+: Small clumps
Hemolysis: Positive reactionComplement fixation
Negative: Sea of red cells
MIXED FIELD REACTIONS
• Serum from group A persons will
agglutinate group B red cells
R E AC T I O N S
• Antibody to B antigens present
(anti-B)
• Serum from group B persons will
agglutinate group A red cells
• Antibody to A antigens present
(anti-A)
• Serum from group O persons will
agglutinate both A and B red cells
R E AC T I O N S
• Antibody to both A and B
antigens present
• Serum from group AB does not
agglutinate either A or B red cells
• Antibody to neither A or B
antigens present
ABO FORWARD GROUPING
Patient’s
RBC’s
1
2
3
4
Reaction
with Anti-A
Negative
+
Negative
+
(SEE CHART)
Reaction with
Anti-B
Negative
Negative
+
+
Interpretation of
Blood Group
O
A
B
AB
+ indicates visual agglutination
ABO REVERSE GROUPING
(SEE CHART)
Patient’s
Serum
Reaction with
A1 Cells
Reaction
with B cells
Interpretation of
Blood Group
1
+
+
O
2
Negative
+
A
3
+
Negative
B
4
Negative
Negative
AB
+ indicates visual agglutination
FREQUENCY OF BLOOD GROUPS
(SEE CHART)
Phenotype
O
White % Black % Mexican
%
45
49
56
Asian %
43
A1
33
19
22
27
A2
8
8
6
Rare
B
10
19
13
25
A1B
3
3
4
5
A2B
1
1
Rare
Rare
• Naturally occurring is misnomer
ABO ANTIBODY
PRODUCTION
• Sources of stimulation found in
nature
• Bacteria
• Seeds from plant
• Pollinating plants
• Foods
ABO ANTIBODIES
• Unique in that it does not require introduction of
foreign red cells by pregnancy or transfusion
• Not present at birth
• Titers not detectable until 3 to 6 months
• Antibody production peaks at 5 to 10 years of age
• Declines with advanced age
• After 65 have low titers that may be undetectable in
reverse typing
OTHER BLOOD GROUPS
• Some blood groups other than ABO can
occasionally cause the production of antibodies
without transfusion or pregnancy
• These antibodies are usually IgM in type
• They are not consistently present in all patients that
have the recessive antigen
TESTING FOR ABO
• Easy to perform
• Lacking the corresponding antigen serves as
confirmation of the forward grouping results
• Very rare (0.01% or random population) to see complete
absence of Anti-A and/or Anti-B in normal, healthy
individuals unless, of course, the person has AB blood
type
• Treacherous if wrong ABO
group is given
• Severe if not fatal
complications
ABO
TYPING
• Forward and Reverse typing
on all patients, note
reciprocal relationship of
antigens and antibody
(discrepancy)
• O is universal donor
• AB is universal recipient
ABO ANTIBODIES
• IgM –
• Cold reacting
• Bind complement (causing lysis)
• Does not cross placenta
• Group A and/or B individuals contain
• Anti-B and/or Anti-A – IgM only (MAJORITY)
• Anti-B and/or Anti-A – IgM and IgG
• Anti-B and/or Anti-A – IgM, and IgA
• Anti-B and/or Anti-A – IgM, IgG and IgA
• IgM is always predominate
• Group O individuals contain
• Anti-A
• Anti –B
• IgG anti-A and anti-B are produced far more
commonly in group O people than in group
A or B people
• Anti-AB
ABO
ANTIBODIES
• Separate cross-reacting antibody that is a
mixture of IgG and IgM or IgG, IgM and IgA
• Crosses the placenta more frequently than
anti-A or anti-B
• Produced in response to exposure to A or B
cells by transfusion or pregnancy
• Knowing the amount of IgG anti-A, anti-B
and anti-AB allows prediction or diagnosis of
hemolytic disease of the newborn (HDN)
caused by ABO incompatibility
INHERITANCE OF ABO BLOOD GROUPS
• Inherit one gene from ABO group from each parent
• Chromosome 9 has one position for the ABO group
• A or B refer to the phenotypes
• What is detectable by testing (EXPRESSED)
• AA, AO, BB, BO, OO denote genotypes
• What genes are on each chromosome (EXPRESSED AND
NON EXPRESSED)
Parent
Phenotypes
Parent
Genotypes
Offspring
Possibility
A XA
AA X AA
AA X AO
AO X AO
A(AA)
A(AA or AO)
A(AA or AO) or O
(OO)
BXB
BB X BB
BB X BO
BO X BO
B (BB)
B(BB OR BO)
B(BB or BO) or O
(OO)
AB O G ROUP S OF T H E OFFSP R I NG FROM
VAR I OUS P OSSI B L E AB O MATI NG S
( SE E CH ART )
Parent
Parent
Phenotypes Genotypes
Offspring Possibility
Pheno & Genotypes
AB X AB
AB X AB
AB(AB) OR A (AA) OR B
(BB)
OXO
OXO
O(OO)
AA X BB
AO X BB
AA X BO
AO X BO
AB(AB)
AB(AB) or B(BO)
AB(AB) or A(AO)
AB(AB) or A(AO) OR
B(BO) OR O (OO)
AXB
AB O G ROUP S OF T H E OFFSP R I NG FROM
VAR I OUS P OSSI B L E AB O MAT I NG S ( SE E
CH ART )
FORMATION OF A, B, H
ANTIGENS
• A, B, and H antigens are not fully developed at birth
• Development begins at 6 weeks of fetal life
• Not fully developed until 2-4 years old
• ABO genes do not actually code for the production of
ABH
• Code for enzymes (glycosyltransferases) that add sugars
to basic precursor substances on the red blood cell
• H gene is inherited independently
from ABO genes
FORMATION OF
A, B, H
ANTIGENS
• H is the backbone on which other
genes are expressed
• H gene is found on Chromosome
19
INTERACTION OF HH AND ABO
GENES
• Inheritance of at least one H gene causes production of
enzyme (alpha-2-L-fucosyl-transferase) that transfers a
sugar
• Genotype HH or Hh is possible (h is an amorph of H)
• Gene is found on Chromosome 19
INTERACTION OF HH AND ABO
GENES
• H gene is known as FUT-1 gene
• 99.9% of population has the H gene
• “h” amorph is very rare – known as the Bombay
phenotype
• Lacks normal expression of ABH antigens even
though person has A and B genes
INTERACTION OF HH AND ABO
GENES
• H substance MUST be formed for other sugars to be
attached to the RBC (regardless to the inherited A or
B gene)
• Bombay people are devoid of all ABO antigens
• Bombay types contain Anti-A, -B, -AB & -H
• They can only be transfused with another Bombay
type
ABO ANTIGENS
1. The blood group is determined by the terminal
sugars on the membranes of their red cells.
2. If neither the A or B gene is present “the O
condition” the antigen is H
3. In the presence of the B gene, the sugar galactose is
added
4. In the presence of the A gene, the sugar N-acetyl
galactosmine is added
TERMINAL SUGAR PLACEMENT
Antigen
Structure
Minimal determinant
structure
H
B
A
*: residue could be glucose in case of glycolipids; yellow shade: minimal determinant or core structure; blue arrow: residue added by
blood group gene product; examples of type 1 and 2 core structures are illustrated above but they can vary widely, as they can be
assembled on at least six possible types of carbohydrate chains; they can reside on a variety of protein or lipid glycan structures
containing branches, repeats, etc.
ABO ANTIGENS
ABO
Grp
Substance
In
Saliva
A
B
H
O
NONE
NONE
++
A
++
NONE
+
B
NONE
++
+
AB
++
++
+
* Non-secretors (sese) will have NO ABH
substances in saliva
FOR MATI ON OF A, B
AND H SOL UB L E
ANT I G E NS
• The presence of ABH soluble
antigens can also be found in all
body secretions IF they inherit
the Se gene.
• Those who inherit the SeSe or
Sese gene will secret these
soluble antigens in body fluids.
If they inherit the sese gene,
they will not be secretors.
1911 discovery of 2 types of A
antigen
A1 and A2 make up 99 percent
of all A groups
ABO
SUBGROUPS
A1 reacts with both Anti-A
and Anti-A1 sera
A2 reacts with only Anti-A
sera
80 % of all Group A (or AB)
are A1 (or A1B)
Note: Anti-A1 sera is NOT
the same as A1 cells
ABO SUBGROUPS
• Antigens present on the RBC surface of A1 and A2 are
presented in two ways:
• Group A1: A and A1 antigens
• Group A2: A antigen only
• Characteristics of A1 & A2 are found in Table 6-14 (page 131)
• Reactivity of anti-H antisera or anti-H lectin with the
different ABO blood groups (figure 6-11)
_____ > _____ > B > ______ > ______ > ______
ADDITIONAL WEAK A
SUBGROUPS
• A3 – demonstrates a mixed field pattern with anti-A
and most of anti-B
• Ax – will not be agglutinated by anti-A reagent but
will agglutinate with anti-AB reagent.
WEAK B SUBGROUPS
• Very rare; less frequent
• Recognized by variations in the strength of reaction using
anti-B and Anti-A,B.
• B3, Bx, Bm and Bel
• See table 6-15 for characteristics of the B Phenotypes
BOMBAY PHENOTYPE
(OH)
• First reported in 1952 in Bombay, India
• Inheritance of a double dose of the “h” gene producing the
very rare genotype hh.
• Causes the ABO genes NOT to be expressed (No H gene =
No place for sugars to attached)
• RBCs are void of normal ABH antigens and fail to react with
anti-A, anti-B and anti-H
• Would phenotype as O blood group
• See Box 6-6 & 6-7 for Bombay Characteristics
ABO DISCREPANCY
• Unexpected reactions in the forward and reverse results
of an ABO blood group typing.
• The forward grouping does not correspond to the results
of the reverse grouping OR
• There is abnormal reactivity present (i.e. mixed field
reaction)
Discrepancies can be
due to technical error
ABO
DISCREPANCY
• Cell suspension too heavy or
too light
• Clerical errors
• Missed hemolysis
• Failure to add reagents
Discrepancies can be
due to other factors.
•
•
•
•
•
Patients age
Diagnosis
Previous transfusion
Medication
Previous pregnancy
ABO DISCREPANCY: 4 GROUPS
• Group 1
• Associated with unexpected reactions in the reverse grouping due to
weakly reacting or missing antibodies
• Group 2
• Associated with unexpected reactions in the forward grouping due to
weakly reacting or missing antigens
• Group 3
• Associated with discrepancies between forward and reverse grouping
caused by protein or plasma abnormalities resulting in rouleaux
formation.
• Group 4
• Associated with discrepancies between forward and reverse grouping
due to miscellaneous problems.
ABO DISCREPANCY – GROUP 1
Causes of Group I Discrepancies
Newborns: Lack antibodies
Elderly: Depressed Antibodies
Resolve Group I
Discrepancies
GOAL: Enhance weak or
missing reaction
Leukemia
1.
Immunosuppressive Drugs
ABO Subgroups
Bone Marrow Transplants &
Plasma Transfusions
Incubate at Room
Temperature for 15 – 30
minutes & centrifuge
2. If there is still no reaction,
then incubate at 4° for 15 –
30 minutes & centrifuge
Always remember to use an
auto and O cell control with
reverse typing
ABO DISCREPANCY – GROUP 2
Causes of Group II Discrepancies
Subgroups of A OR B
Leukemia weakened A or B
antigens
Hodgkin’s disease
Resolve Group II
Discrepancies
GOAL: Enhance
weak or missing
reaction
1. Incubate at
Room
Temperature for
15 – 30 minutes &
centrifuge
2. If there is still
no reaction, then
incubate at 4° for
15 – 30 minutes &
centrifuge
“Acquired B” phenomenon
3. RBC’s can be treated with enzymes and
retested with antisera. In the case of
suspected “Acquired B”, treat RBC’s with
acetic anhydride reacetylates and retest.
ABO DISCREPANCY – GROUP 3
Causes of Group III Discrepancies
Elevated levels of globulin-multiple
myeloma, Waldenstroms macroglobinemia
Elevated levels of fibrinogen
Plasma expanders such as dextran and
polyvinylpyrrolidone
Resolve Group III
Discrepancies
GOAL: Remove
interfering substance
1. Perform a saline
dilution or saline
replacement to free
cells
Wharton’s jelly in cord blood samples
2. Wash cells 6 – 8
times with saline to
alleviate the rouleaux
due to Wharton’s jelly
ABO DISCREPANCY – GROUP 4
Causes of Group IV Discrepancies
Resolve Group IV Discrepancies
Cold reactive autoantibodies
GOAL: Eliminate spontaneous agglutination
Circulating RBC’s of more than one
ABO group due to transfusion or
transplant
1. Forward: Incubate at 37° for a short time and
then wash at 37° 3 times and retype
Unexpected ABO isoagglutinins
Unexpected non-ABO
alloantibodies
2. Reverse: Warm to 37° mix and test at 37°
* If forward doesn’t resolve with 1, then treat patient’s cells with
DTT.
* If reverse doesn’t resolve with 2, try an autoabsorbtion from the
serum and then use the absorbed serum to repeat the typing at
room temperature .
HOMEWORK
• CHAPTER 6
• Review ABO discrepancies and case study
(PGS. 136-145)
• Review ABO Discrepancy Tutorial online
and case studies