The document summarizes the history and science behind blood grouping and the ABO blood group system. It describes how Karl Landsteiner discovered the major ABO blood groups in 1901. It explains the antigens and antibodies present in each blood group according to Landsteiner's rule. The genetics and biochemistry of the ABO blood group system are covered, including how the H, A, and B antigens are synthesized on red blood cells. Common blood grouping techniques like forward and reverse grouping are also summarized.

1. BloodGrouping

2. History - Karl
Landsteiner
 Discovered the ABO Blood
Group System in 1901
 He and his five co-workers
began mixing each others red
cells and serum together and
inadvertently performed the
first forward and reverse ABO
groupings

3. Landsteiners
Rule
 If an antigen (Ag) is present on a patients red blood
cells the corresponding antibody (Ab) will NOT be
present in the patients plasma, under ‘normal
conditions’.

4. MajorABO
BloodGroup
ABO
Group
Antigen
Present
Antigen
Missing
Antibody
Present
A A B Anti-B
B B A Anti-A
O None A and B Anti-A&B
AB A and B None None

5. ABO Basics
 Blood group antigens are actually sugars attached to
the red blood cell.
 Antigens are “built” onto the red cell.
 Individuals inherit a gene which codes for specific
sugar(s) to be added to the red cell.
 The type of sugar added determines the blood group

6. Principle of
blood
grouping
There are two principles
1-almost all normal healthy individuals above 3-6
months of age have “ naturally occurring Abs” to
the ABO Ags that they lack
These Abs termed naturally occurring because they
were thought to arise without antigenic stimulation

7. Principle of
blood
grouping
2- These “naturally occurring” Abs are mostly IgM
class. That means that, they are Abs capable of
agglutinating saline/ low protein suspended red
cell without enhancement and may activate
complement cascade.

8. ABO and H
Antigen
Genetics
 Genes at three separate loci control the occurrence and
location of ABO antigens
 The presence or absence of the A, B, and H antigens is
controlled by the H and ABO genes

9. Location
 The presence or absence of the ABH antigens on the
red blood cell membrane is controlled by the H gene
 The presence or absence of the ABH antigens in
secretions is indirectly controlled by the Se gene

10. HAntigen
 The H gene codes for an enzyme that adds the sugar
fucose to the terminal sugar of a precursor substance
(PS)
 The precursor substance (proteins and lipids) is formed
on an oligosaccharide chain (the basic structure)

11. RBC Precursor
Structure Glucose
Galactose
N-acetylglucosamine
Galactose
Precursor
Substance
(stays the
same)
RBC

12. Formation of
the H antigen Glucose
Galactose
N-acetylglucosamine
Galactose
Precursor
Substance
(stays the
same)
RBC
H antigen
Fucose

13. H antigen
 The H antigen is the foundation upon which A and B
antigens are built
 A and B genes code for enzymes that add an
immunodominant sugar to the H antigen
 Immunodominant sugars are present at the terminal ends of
the chains and confer the ABO antigen specificity

14. A and B
Antigen
 The “A” gene codes for an enzyme (transferase) that
adds N-acetylgalactosamine to the terminal sugar of
the H antigen
 N-acetylgalactosaminyltransferase
 The “B” gene codes for an enzyme that adds D-
galactose to the terminal sugar of the H antigen
 D-galactosyltransferase

15. Formation of
theA antigen Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose
N-acetylgalactosamine
A antigen

16. Formation of
the B antigen Glucose
Galactose
N-acetylglucosamine
Galactose
RBC
Fucose
Galactose B antigen

17. H antigen
 Certain blood types possess more H antigen than
others:
O>A2>B>A2B>A1>A1B

18. Why do Group
O individuals
have more H
antigen than
the other
groups?
Group O
individuals have
no A or B genes
to convert the H
antigen to A or B
antigens….that
means more H
antigen sites

19. Group O Group A
Many H
antigen sites
Fewer
H antigen
sites
A
A A
AA
Most of the H antigen sites in a
Group A individual have been
converted to the A antigen

20. Genetics
 The H antigen is found on the RBC when you have the
Hh or HH genotype, but NOT from the hh genotype
 The A antigen is found on the RBC when you have the
Hh, HH, and A/A, A/O, orA/B genotypes
 The B antigen is found on the RBC when you have the
Hh, HH, and B/B, B/O, orA/B genotypes

21. Bombay
Phenotype
(Oh)
 Inheritance of hh
 The h gene is an amorph and results in little or no
production of L-fucosyltransferase
 Originally found in Bombay
 Very rare (130 worldwide)

22. Bombay
Phenotype
(Oh)
 The hh causes NO H antigen to be produced
 Results in RBCs with no H, A, or B antigen (patient
types as O)
 Bombay RBCs are NOT agglutinated with anti-A, anti-
B, or anti-H (no antigens present)
 Bombay serum has strong anti-A, anti-B and anti-
H, agglutinating ALL ABO blood groups
 What bloodABO blood group would you use to
transfuse this patient??
Another Bombay
Group O RBCs cannot be given because they still
have the H antigen
You have to transfuse the patient with blood that
contains NO H antigen

23. ABO
antibodies
 GroupA serum contains anti-B
 Group B serum contains anti-A
 GroupAB serum contains no antibodies
 GroupO serum contains anti-A, anti-B, and anti-A,B

24. ABO
antibodies
 IgM is the predominant antibody in GroupA and Group
B individuals
 Anti-A
 Anti-B
 IgG (with some IgM) is the predominant antibody in
GroupO individuals
 Anti-A,B (with some anti-A and anti-B)

25. ABO
antibodies
 Reactions phase: Room temperature
 Complement can be activated with ABO antibodies
(mostly IgM, some IgG)
 High titer: react strongly (4+)
 Usually present within the first 3-6 months of life
 Stable by ages 5-6 years
 Decline in older age
 Newborns may passively acquire maternal antibodies
(IgG crosses placenta)
 Reverse grouping (with serum) should not be performed on
newborns or cord blood

26. ABO
routine testing
 Several methods for testing the ABO group of an
individual exist.The most common method is:
 Serology: This is a direct detection of the ABO
antigens. It is the main method used in blood
transfusion centres and hospital blood banks.
 This form of testing involves two components:
a)Antibodies that are specific at detecting a particular
ABO antigen on RBCs.
 b) Cells that are of a known ABO group that are
agglutinated by the naturally occurring antibodies in
the person's serum.

27. ABO
ROUTINE
TESTING
DIRECT OR FORWARD GROUPING
Test for antigens
• Patient’s cells containing unknown antigens tested
with known antisera
• Antisera manufactured from human sera
Aantisera used:
Antisera Color Source
Anti-A Blue Group B donor
Anti-B Yellow Group A donor
Anti-A,B Red Group O donor

28. Forward
Grouping
 Reaction of patient red blood cells tested with
Reagent anti-A and anti-B antisera
 Slide: 20-40% RBC suspension + anti-serum
 Tube (12x75mm): 2-5% RBC suspension + anti-serum
(centrifuge before read)

29. Forward
Grouping
Reaction Patterns for ABO Groups
Blood group Agglutination with
Anti-A
Agglutination with
Anti-B
A + -
B - +
AB + +
O - -

30. Reverse
grouping
• serum is combined with cells having known Ag
content in a 2:1 ratio
• uses commercially prepared reagents containing
saline-suspended A1 and B cells

31. Reverse
grouping
Reaction Patterns for ABO Groups
Blood Group Agglutination with
A cells
Agglutination with
B cells
A - +
B + -
AB - -
O + +

32.  Grading of Agglutination:
Negative (0) No clumps or aggregates
Weak (+/-) Tiny clumps or aggregates barely
visible macroscopically or to the
naked eye
1+ Few small aggregates visible
macroscopically
2+ Medium-sized aggregates
3+ Several large aggregates
4+ One solid aggregate

33. ABO blood
group
(forward blood
grouping)
Patient Red CellsTested With
InterpretationAnti-BAnti-APatient
001
04+2
4+03
4+4+4

34. ABO blood
group
(forward blood
grouping)
Patient Red CellsTested With
InterpretationAnti-BAnti-APatient
O001
A04+2
B4+03
AB4+4+4

35. Reverse
Grouping
(Confirmatory
grouping
Patient SERUMTestedWith
InterpretationB CellsA1 CellsPatient
4+4+1
4+02
04+3
004

36. Reverse
Grouping
(Confirmatory
grouping
Patient SERUMTestedWith
InterpretationB CellsA1 CellsPatient
O4+4+1
A4+02
B04+3
AB004

37. Forward &
reverse
ABO blood
grouping
Reaction of CellsTested With
Reaction of SerumTested
Against ABO
Group
Anti-A Anti-B A1 Cells B Cells
1 0 0 + + O
2 + 0 0 + A
3 0 + + 0 B
4 + + 0 0 AB

38. Forward &
reverse
ABO blood
grouping
Reaction of CellsTested With
Reaction of SerumTested
Against ABO
Group
Anti-A Anti-B A1 Cells B Cells
1 0 0 + +
2 + 0 0 +
3 0 + + 0
4 + + 0 0

39. ID card
system
 This ID-Card contains a mixture of human polyclonal
and monoclonal anti-A, human polyclonal anti-B and
human polyclonal anti-D antibodies.
 The microtube ctl is the negative control.Two
microtubes with neutral gel serve for reverse grouping
with A1 and B cells.

40.  Fully-automatic walk-away for ID-Cards
 Stand alone instrument
 Continuous sample loading
 Continuous reagent loading
 Priority samples (STAT)
 Reagent stock
 High throughput

41.  Optimized for small blood volumes
 Dispense verification
 Easy-to-use
 Full test menu
 Wi-Fi
 Touchscreen 17"
 Host connectivity
 Internal & external validation
 Capacity
180 samples
240 ID-Cards
28 reagent vials

42. Thank you….