1682 William Harvey Discovered Blood Circulation

1658 Jan Observed and described Red Blood

Swammerdam Cells
1665 Richard Lower First recorded successful blood
transfusion in dogs
1667 Jean Baptiste Separate reports on successful blood
Denis & Richard transfusions from sheep to humans
OUTLINE Lower
I. Introduction 1818 James Blundell First Successful Transfusion of
 Immunohematology Human Blood
1869 Braxton Hicks Recommended sodium Phosphate as
 History of Transfusion a non-toxic anticoagulant
II. Immunologic Principles 1873 US physicians transfuse milk from
III. ABO Blood Group cows, goats and humans
1884 Saline infusion replaces milk as a
 ABO Laboratory Tests blood substitute due to increased
 ABO Subgroups frequency of adverse reactions to milk
 Bombay Phenotype 1900 Karl Landsteiner ABO Blood Group System
 ABO Discrepancies 1907 Ludvig Hektoen Introduction of crossmatching
between donors and patients
1912 Rueben First to perform blood transfusion
INTRODUCTION Ottenberg using blood typing and
IMMUNOHEMATOLOGY crossmatching
- Merges aspects of Hematology, Immunology, and 1914 Roger Lee Packed RBC Products
Group O as universal donor
Genetics. Group AB as universal recipient
- Serologic, genetic, biochemical and molecular study
of antigens associated with membrane structures on Plasma Products
Group O as universal recipient
the cellular constituents of the blood. Group AB as universal donor
- Immunologic reactions involving all blood components
and constituents. 1914 Long-term anticoagulants were
developed for longer blood
 Blood Banking - process of collecting, separating and
preservation (e.g. Sodium Citrate)
storing of blood. Richard Lewisohn Determined the minimum amount of
1915
 Blood Transfusion– administration of blood and blood citrate needed for anticoagulant
products. (63mL)
1916 Francis Rous Citrate dextrose solution for blood
 Bloodletting – one of the oldest medical practice and is J.R. Turner preservation
linked to many cultures which were believed to cure 1939 Alex Wiener RH Blood group system
sickness. Karl Landsteiner
Philip Levine
 Galen of Pergamon – discovered that arteries are filled R.E. Stetson
with blood and not air. Also,, believes that blood does not 1941 Isodor Radvin Treated victims of the Pearl Harbor
circulate hence remain motionless. attack with Cohn’s albumin for shock
 1943 J.F. Loutit Introduction of acid citrate dextrose
Patrick Mollison
HUMORISM Coombs Use of AHG to identify “incomplete”
1945
- Balance of four humors of the body; Phlegm, Blood, Mourant antibodies
Yellow Bile, and Black Bile. Race
1950 Audrey Smith Use of glycerol cryoprotectant for
- Any excess/deficiency of these bodily fluids directly freezing red blood cells.
influences the temperature and health of a person. 1950 Carl Walter Use of plastic blood bags for collection
W.P. Murphy
1961 Recognizing the role of platelets in
reducing mortality from hemorrhage
among cancer patients
1962 First AHF concentrate was developed
through fractionation
1964 Plasmapheresis was introduced as a
means of collecting plasma for
fractionation
1967 RH immunoglobulin was commercially
introduced
1971 HBsAg testing for donated blood
1972 Apheresis was used to extract one
cellular component
1979 CPDA-1 was developed allowing
HISTORY OF TRANSFUSION PRACTICES longer storage of red blood cells
1981 First AIDS case was reported
YEA CONTRIBUTION
1983 Additive solutions used extending
R shelf life of RBCs further
1492 Pope Innocent Blood was taken from 3 young boys
1984 HIV was identified as the causative
VII/VIII and given to the Pope through mouth agent of AIDS
in hopes to cure his illness Testing of donor blood for HIV-1 and
1992
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 HIV-2 antibodies was implemented
2002 Nucleic acid amplification test for HIV  IgG - a smaller molecule, with a pair each of heavy and
and HCV was licensed by the US FDA
light chains of amino acids. Can just coat but not
agglutinate the cells.
IMMUNOLOGIC PRINCIPLES
 IgM – has 5 pairs joined together by J chain. Can
- Primary immunological components; antigen and
agglutinate the cells bearing corresponding antigen.
antibodies provides basis for blood bank testing and
reactions.
IgG are more reactive in WARM temperature
- Immunologic reaction in immunology involves all
IgM are more reactive in COLD temperature
blood components and its constituents including your
plasma, red blood cell, platelets, and white blood cell
IgG is subdivided to 4 classes; IgG1, IgG2, IgG3, IgG4
- We have 2 Major Reactants in primary immunological
*All can cross placenta except IgG2
reaction; the antigen and antibody. This provides the
basis of blood bank testing and reactions such as Rh
- Small quantities of IgG are present in group A and B
typing, ABO typing and Cross matching.
individuals
- Anti-A, B – cross reacting IgG antibody are found in
 CARDINAL RULE IN BLOOD BANK:
the sera of group O individuals
- The antigens are found on the surface of Red blood
- This produces strong agglutination during ABO testing
cells and the antibodies are found in Serum or
- Reacts preferentially at room temperature (20-24C) or
Plasma.
below and activates at 37C
- Production is initiated at birth, becomes detectable at
3-6 months
TAKE NOTE:
- It causes rapid intravascular hemolysis and can cause
- ANTIGENS --- RBC
death of the patient
- ANTIBODIES --- SERUM / PLASMA
ABO BLOOD GROUP SYSTEM
 MAJOR CHARACTERISTICS OF THE ANTIGEN - Single most important blood group for the selection
(In order for antigens to be immunogenic) and transfusion of blood.
- Widely expressed in tissues and body fluids including
1.) Chemical Structure of Antigen
red blood cells, platelets and endothelial cells
- It is said that if composed mainly of oligosaccharide - Transfusion of incompatible ABO type result to almost
then it tends to stimulate the ABO antibody immediate lysis of donor RBCs
production, while those antigens containing protein
 Three Antigens : A, B, H
are able to stimulate IgG antibodies.
 Two Major Antibodies : Anti-A and Anti-B

2.) Degree of Foreignness

TYPE ANTIGEN ANTIBODY
- Chemical structure that determines the
A A Anti – B
immunogenicity and antigenicity of antigen.
- The greater the degree of antigenic determinant is B B Anti – A
recognized as non-self by individual immune system AB A,B -
the more antigenic it is. O - Anti-A , Anti-B
- The More Foreign = The More Antigenic
INHERITANCE
3.) Number of Antigen - Described by Bernstein ion 1924
- Amount of antigen introduced in the body to elicit - Individuals inherit one ABO gene from each parent
antibody production and these two genes determine the ABO blood group.
- The Higher Dose = The Higher the Antibody - Follows simple mendelian genetics.
Production - ABO gene is found in chromosome 9 (A, B, or O
gene); codominantly expressed
4.) Route of Administration - Phenotypes: A, B, AB, O
- Intraperitoneal offers more stronger stimulus of - Genotypes: AA, AO, BB, BO, AB, OO
immune response to produce antibody than the - “A and B genes produce detectable product while O
subcutaneous or intramuscular route gene is amorphic or silent so it does not produce
detectable antigen or product”
 IMMUNOGLOBULINS
- Five types  NOTE:
IgG-IgM-IgA-IgD-IgE A , B , AB – Autosomal Dominant
- Blood group antibodies are mainly confined to IgG O – Autosomal Recessive
and IgM *Dominant means possible expression of product of
particular Gene.

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 ABH ANTIGENS LABORATORY TESTS FOR ABO
- Results from the interaction of these three distinct FORWARD TYPING/GROUPING
genes; ABO, H, Se. - Using known sources of antisera to detect antigens
- Codes for the production of specific on an individual’s RBC
glycosyltransferase that add sugar to a basic - Slide or tube method are used
precursor substance. - Steps in forward typing :
- A, B, and H antigens – formed in basic precursor 1. Blood samples are taken then RBCs are
substance. separated from the plasma/serum via
centrifugation.
 Type 1 - precursor of ABH soluble substances 2. Mix the RBCs with reagents (anti-A, anti-B
- terminal galactose is attached to GLNAC antibodies)
in a β 1-3 linkage 3. Wait for agglutination
- in secretion (saliva,tears) and in plasma - Results :
- type 1 chains are converted into H antigen  A antigen containing RBCs will agglutinate in
by enzymatic action (fucosylation) of Se tube with Anti-A antibodies
gene product  B antigen containing RBCs will agglutinate in
 Type 2 - precursor of ABH antigens on red cell tube with Anti-B antibodies
membrane  If it contains BOTH A and B antigen RBCs
- terminal galactose is attached to GLNAC then it will agglutinate on both tubes that has
in a β 1-4 linkage Anti-A and Anti-B antibodies
- on the surface of RBC  If it contains NEITHER A or B antigen RBCs
- type 2 chains are converted to H antigen then it will have no agglutination reaction on both
by enzymatic action (fucosylation) of H tubes
gene product

 Immunodominant Sugar – sugar that occupies the

terminal portion of the precursor chain and confers
blood group specificity

- Reagents of ABO Antisera (Lectins)

 Anti-A
- Trypan blue
 Anti-B
- Acriflavine yellow
ABH SOLUBLE SUBSTANCE
 Anti-H
- Found in all body secretions
- Ulex europaeus
- Presence is dependent on ABO gene and SeSe/Sese
gene (secretor gene)
REVERSE TYPING/GROUPING
- H soluble antigen can be modified to A and B soluble
- To detect ABO antibodies in the serum of an
antigen.
individual by using known RBCs reagents (A and B
ABO Group A soluble B soluble H soluble
cells)
A ++ - +
B - ++ + - Steps in reverse typing :
O - - ++ 1. Blood samples are taken and serum/plasma are
AB ++ ++ + separated from the RBCs via centrifugation
2. The Serum/Plasma of the individual is then mixed
NOTE: with the RBCs reagents of known A and B
- The role of se gene directly controls the presence antigen.
and absence of A, B, and H antigen in the 3. Wait for agglutination.
secretion. - Results :
- Inheritance of A and B genes -> results in the  Serum with Type B or O that has Anti-A
expression of A and B gene products (antigens) on antibodies will agglutinate to A cells
erythrocytes, but are NOT the direct products of ABH  Serum with Type A or O that has Anti-B
genes antibodies will agglutinate to B cells
 Serum with Type O that has BOTH Anti-A and
Anti-B antibodies will agglutinate to BOTH A
and B cells

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  Serum with Type A and B that has NEITHER - Newborn – ABO antibodies not detectable until 3-6
antibodies will not agglutinate to NEITHER A months of age
nor B cells - Elderly patients – production of ABO antibodies is
depressed
- Leukemic patients – hypogammaglobulinemia
- Patients using immunosuppressive drugs –
hypogammaglobulinemia
- Bone marrow transplant – hypogammaglobulinemia
- Patients in plasma transfusion/exchange – diluting
effect
- Chimerism – more than one ABO in an individual

 GROUP II
- Unexpected reactions in the forward grouping due to
weak or missing ANTIGENS.
- May be due to: ABO subgroups, leukemia, Hodgkin’s
disease, acquired B phenomenon, BGSS, chimerism
ABO SUBGROUPS - Blood Group-specific Soluble Substance (BGSS) –
- The subgroups of A and B are caused by decreased seen in certain disease such as stomach and
amounts of antigens on the red blood cells. They are pancreatic cancer
inherited conditions. - Excess BGSS will neutralize reagent anti-A
and anti-B, causing a weak or false negative
- The most common are subgroups of A. result.
Approximately 80% of the A’s and AB’s have normal
expressions of A1. Most of the other 20% are either  GROUP III
A2 or A2B. - Discrepancies between forward and reverse grouping
- Both subgroups are equally strong in Anti-A due to protein or plasma protein abnormality resulting
forwarding group. Anti-A1 antibody is specifically to rouleaux formation or pseudo agglutination.
reacts to A1 but not A2. - Elevated globulin levels, elevated fibrinogen, plasma
expanders, Wharton’s jelly in cord blood sample
BOMBAY PHENOTYPE - Resolution: washed RBC’s with NSS
- Inheritance of double dose of the h gene producing hh
 GROUP IV
genotype (Oh)
- Discrepancies between forward and reverse grouping
- The Bombay Blood group lacks H gene and therefore
due to miscellaneous problems
cannot make H antigen on RBCs. - Resolution: perform antibody panel
- Since H antigen is the precursor for the A and B
antigens, these antigens also are not made.
- Bombay RBCs will fail to react to anti-H lectin.
- Type O donor cells + Oh recipient -> immediate lysis
of donor red cells
- The only cells Bombay individuals do not agglutinate
are those from other Bombay blood individuals since
they also lack H antigen.

ABO DISCREPANCIES
- Unexpected reaction in the forward and reverse
typing/grouping.
- Wash cells with saline 3-4x and repeat all tests and
test for antibodies
- Test for subgroups of A using anti-A and anti-A1
- Use cell panels to detect specificity of abnormal
antibodies
- Reverse typing: usually resolved by repeating test on
the same sample using saline suspension.
- Any technical factor that may have given ride to the
ABO discrepancy should be reviewed and corrected

 GROUP I
- Unexpected reactions in the reverse grouping due to
weak or missing ANTIBODIES.
- Agglutination reactions with ABO antibodies should
be very strong (4+)

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 FISHER-RACE: DCE TERMINOLOGY
- Three alleles: D/d, C/c, and E/e
- Five antigens: D. C. E. c. e
- D -> no D locus -> no antigenic products
- Each gene and corresponding antigens were given
the same letter designation
OUTLINE
- Individuals inherit a set of Rh gene from each parent
I. Rh Blood Group System
- Co-dominantly expressed
 History
II. Fisher Race: DCE Terminology
WIENER: RH-hr TERMINOLOGY
III. Wiener: Rh-hr Terminology
- Postulated that Rh gene produces an agglutinogen
IV. Rosenfield: Alpha-Numeric Terminology
that contains a series of blood factors.
V. ISBT Committee: Updated Numeric Terminology
VI. Genetics
Wiener to Fisher-Race Translation
VII. D Antigen
 Weak D Phenotypes
 Genetic Weak D
 C Trans Effect
 Partial D or D Mosaic
VIII. LW Antigens
IX. Rh Antibodies
 Rh-associated Hemolytic transfusion reaction
 Rh-associated Hemolytic disease of the
newborn
 ABO HDN ROSENFIELD: ALPHA-NUMERIC TERMINOLOGY
X. Anti-D Antisera - Numerical system; Rh1 to Rh5
- Demonstrates the presence or absence of the antigen
RH BLOOD GROUP SYSTEM on RBCs
- Second most important human blood group system - Minus sign designates the absence of the antigen
- Currently composed of 61 different antigens
- Antibodies are immune in nature; causes significant
HDFN (Hemolytic Disease of the Fetus and Newborn)
and HTR (Hemolytic Transfusion Reaction).
- Rh typing – tests for the presence of D antigen - Example:
- When they agglutinate they are Rh positive and if they D- : Rh1-
didn’t agglutinate they are Rh negative C+ : Rh2
E+ : Rh3
HISTORY c+ : Rh4
- 1939 Levine and Stetson defined D antigen (Rh e- : Rh5-
Factor)
- 1940 Landsteiner and Weiner discovered anti-Rh ISBT COMMITTEE: UPDATED NUMERIC TERMINOLOGY
(named after Rhesus monkey) - Adopts a six-digit number for each blood group
- Agglutinated 85% human RBCs specificity.
- First 3 digits = system; last 3 digits = antigen
- 15% RBCs did not agglutinate
- Rh = 004
- The antibody was similar to guinea pig and rabbit - D = 001, C = 002, E = 003, c = 004, e = 005
antibodies produced when stimulated with rhesus - Phenotype designation is similar with Rosenfield
monkey RBCs
- Although they were later shown to be different, the GENETICS
name remained - Two closely linked genes control the expression of
ALL Rh antigens (condominant alleles)
NOMENCLATURES  RHD gene – determines the expression of
- Fisher-race - Wiener the D antigen
- Rosenfield - ISBT  RHCE gene – determines the expression of
the C. c. E. and e antigens
 Basis: - Found in chromosome 1
- Postulated genetic mechanisms - Inherited as codominant alleles; one haplotype from
- Presence or absence of a given antigen each parent
- Combined efforts of ISBT Committee on Terminology - RHAG
for Red Cell Surface Antigens
 Rh-associated glycoproteins; found in
chromosome 6
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  Must be present for proper expression of Rh - If the patient is transfused with D positive red cells,,
antigens they may develop an anti-D alloantibody to the part
of antigen (epitope) that is missing.

D ANTIGEN LW ANTIGENS
- The most important red cell antigen in blood banking - Phenotypically similar to Rh.
because the antibody in the Rh can cause transfusion - Anti-LW reacts strongly with most D-positive RBCs,
reactions and hemolytic disease of the newborn weakly with D-negative RBCs, and no reaction with
Rh null RBCs.
- The D antigen of your Rh system can elicit/stimulate
 Rh null – no Rh antigens being expressed on
the production of D antibody. the surface of RBCs.
- Most immunogenic (D>c>E>C>e) - Rh null condition occurs when the red cells have no
- Rh-positive – indicates the presence of D antigen Rh antigens sites
(or weak D) - The lack of antigens causes the red cell membrane to
- Any Rh-negative result must be confined by weak D appear abnormal leading to:
testing  Stomatocytosis
- Phases of weak D testing:  Hemolytic anemia
- Two Rh null phenotypes:
1. 37C incubation
1. Regular type – gene inherited, but not expressed
2. Indirect AHG testing 2. Amorph type – RHD gene is absent, no
- Interpretation: expresion of RHCE gene
1. Rh-positive – agglutination in any phases of - Three LW alleles:
testing 1. LWa – common
2. Rh-negative – no agglutination in all phases 2. LWb – rare
3. LW – amorph
WEAK D PHENOTYPE
- Some D-positive RBCs DO NOT react at immediate AABB STANDARDS
spin using commercial anti-D - Requires weak D testing on all donor red blood cells
- In these cases AHG testing is needed to determine that do not agglutinate at immediate spin
the D status - DO NOT require weak D testing on recipient blood
- Weak D can be inherited in three ways:
 Incomplete/Partial antigen (D mosaic) NOTE:
 Due to the position effect (Deletion) - AABB Standards stands for American Association of
 Weakened expression of D antigen Blood Bank Standards
- ALL DONOR RED CELLS that DO NOT
GENETIC WEAK D AGGLUTINATE at the immediate spin REQUIRES
- Inheritance of Rh genes that code for a weakened WEAK TESTING.
expression of D antigen
- Antigen is complete but the number is few RH ANTIBODIES
- Not naturally occurring
C TRANS EFFECT / DELETION - Produced as a result of immune stimulation from
- Also known as position effect or gene interaction exposure to the antigen through transfusion and/or
effect. pregnancy
- D gene in trans position to the allele carrying C gene - Implicated in transfusion reactions
- Antigen is normal and complete but steric - Because they are IgG, the can cross placenta and
arrangement of C in relation to D appears to interfere can cause Hemolytic Disease of the Newborn (HDN)
with the expression of D. - IgG1 and IgG3 Anti D – most clinically significant
- No reaction when RBCs are tested with anti-E, anti-e, (rapidly cleared)
anti-C, or anti-c  IgG1 – best placental crosser
- Requires transfusion of the other D-deletion red cells  IgG3 – can activate the complement proteins
because these individuals may produce antibodies - Do not bind complement proteins
with single or separate specificities - RBCs sensitized with the anti-D destroyed through
- Written as D- or –D- extravascular hemolysis
*For example; the patient is a really weak D due to - Most common cause of HDFN/Erythroblastosis fetalis
the D-deletion you have to write D-(D negative).
Meaning there is no reaction. NOTE:
- ABO is naturally occurring
PARTIAL D / D MOSAIC - Rh antibodies are NOT naturally occurring
- One or more epitopes within the entire antigen is - Rh antibodies => HDN (Erythroblastosis fetalis)
missing or altered.
- Alloantibody against the missing epitope may be
produced upon exposure to the complete D antigen.
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RH-ASSOCIATED HEMOLYTIC TRANSFUSION REACTION - ABO HDN can occur during First pregnancy
- Circulating anti-D appears within 20 days post- because prior sensitization is not necessary
primary exposure, and 2-7 days post-secondary - ABO HDN is less severe than Rh HDN because there
exposure. are less RBC destruction
- Results in extravascular hemolysis of IgG-coated
 Fetal RBCs are less developed at birth so
RBCs.
- Clinical manifestations: there is less destruction by maternal
1. Unexplained fever antibodies.
2. Elevated serum bilirubin  When delivered, infants may present with
3. Decreased hemoglobin and haptoglobin anemia or normal hemoglobin levels
4. Positive direct AHG test – in vivo attachment of
antibody to RBC antigens
ANTI-D ANTISERA
RH-ASSOCIATED HEMOLYTIC DISEASE OF THE
NEWBORN (HDN)
- It is destruction of RBCs of the fetus and the newborn
by antibodies produced by the mother
- Only IgG antibodies are involved because it can
cross the placenta
- Most severe form of HDN
- 33%% of HDN is caused by Rh incompatibility
- Sensitization usually occurs very late in pregnancy,
so first Rh-positive child is not affected
- Bleeds most often occur at delivery False Positive Reactions with Rh Typing
- Mother is sensitized
- Subsequent offspring that are D-positive will be
affected

Pathophysiology of HDN

- Most often involves antigens of the Rh and ABO

blood group system,, but can result from any blood
False Negative Reactions with Rh Typing
group.
- Remember: the fetus is POSITIVE for an antigen and
the mother is NEGATIVE for the same antigen

NOTE:
- Rh incompatibility, remember the HDN occur when
the FETUS is POSITIVE for the ANTIGEN, and the
antigen of the fetus will stimulate the production of
MATERNAL ANTIBODIES.
- These maternal antibodies will destroy the fetal
RBCs.

- Rh-immune globulin:
 Purified preparation of IgG anti-D
 Given during pregnancy and following
delivery of a D-positive fetus
 These are given to avoid alloimmunization

ABO HEMOLYTIC DISEASE OF THE NEWBORN (HDN)

- most common cause of HDN but are less severe
- 60% of HDN are due to ABO incompatibility
- Usually the Mother is type O and the Child has the
A or B antigen
- Group O individuals have high titer of IgG anti-A and
anti_B in addition to having IgM anti-A and anti-B

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 - Can be neutralized by Lewis’s antigens in plasma or
saliva.

OUTLINE
I. Lewis System
 Lewis Gene
 A B and Leᵇ soluble substance
II. Lewis Phenotypes
III. Lewis Antibodies CLINICAL SIGNIFICANCE OF LEWIS ANTIBODIES
- Generally considered insignificant because:
LEWIS SYSTEM 1. They can be neutralized by Lewis antigens in the
- Only antigen system that is not produced by red blood plasma
cells. 2. Antigens dissociate from RBCs are readily as
- Produced by tissue cells and secreted into bodily they bind to it.
fluids (antigens are present in secretions and 3. They are generally IgM and cannot cross the
plasma). placenta and cause HDFN and HTR.
- Antigens are adsorbed onto red blood cell membrane 4. Presence of Lewis antibodies in the patient’s
from plasma. serum does not require transfusion of Le-
- Antigen expression depends on the interaction of the negative RBCs (as long as 37C and AHG phase
following genes: are compatible).
 Le gene
 Se gene
 ABO gene

LEWIS GENE
- Found in chromosome 19
- Distantly linked to Se and H gene.
- Codes of L-fucosyltransferase that adds L-fucose to
type 1 precursor substance.
- Enzyme competes with Se and ABO gene by
attaching L-fucose to C4 of N-acetylglucosamine of
type 1 precursor chain.

A, B, AND LeᵇSOLUBLE SUBSTANCE

- A and B soluble substance can serve as a substrate
for the Lewis enzyme, forming ALeᵇ and Bleb
antigens.
- Le⁹ and Leᵇ are no longer substrates for ABO and Se
enzymes.

LEWIS PHENOTYPES

LEWIS ANTIBODIES
- Frequently detected in antibody screening.
- Considered naturally occurring and are generally
produced by Le(a-b-) individuals; occur frequently in
pregnant women.
- IgM (can cause occasional in vivo and in vitro
hemolysis); increased reaction with enzyme-treated
cells.
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 - In rare cases, IgG anti-M can cause hemolytic
disease of the newborn or hemolytic transfusion
reaction
- Clinically significant if it is reactive at 37C

ANTI-N
OUTLINE
- Cold-reactive IgM or IgG saline agglutinin
OTHER MAJOR BLOOD GROUP SYSTEM
- Does not bind complement; does not react with
I. MNS Blood Group System enzyme-treated cells
II. P Blood Group System - Demonstrate dosage effect; clinically significant
III. I Blood Group unless reactive at 37C
IV. Kell Blood Group System - In rare cases, may cause Hemolytic disease of the
V. Duffy Blood Group System newborn
VI. Kidd Blood Group System - Also seen in renal patients who underwent dialysis
VII. Lutheran Blood Group System using formaldehyde (anti-N) which alter M and N
antigens thus recognized as foreign
MNS BLOOD GROUP SYSTEM
- Composed of 43 antigens; Anti-M and Anti-N were ANTI-S AND ANTI-s
reported (1972). - IgG, reactive at 37C and AHG phase
- Landsteiner and Levine immunized rabbits with - Demonstrates dosage effect
human RBCs. - May or may not react with enzymes-treated cells
- M and N – antithetical antigens. - More significant compared to anti-M and anti-N
- Walsh and Montgomery (1947) – discovered S - May bind complement and have been implicated in
antigen severe HTR and HDN.
- 1951 – s antigen was discovered
- Wiener (1953) – U antigen was discovered P BLOOD GROUP SYSTEM
- Located at chromosome 4 - Discovered in 1972 by Landsteiner and Levine when
- GYPA and GYPB genes – codes for GPA and GPB they injected human RBCs to rabbits which then
- Codominant expression (both alleles are expressed, produced anti-P
neither allele is dominant or recessive) - 1951, Levine described anti-Tj⁹ (anti-P,P1,Pk) are
produced by P null individuals
M AND N ANTIGENS - Involves antigens P, P1, Pk
- Found on glycophorin A (GPA) – major sialic acid-rich - Cannot be considered a single blood group
glycoprotein (sialoglycoprotein).  P1 = blood group
- Well developed at birth  P = globoside blood group
- Easily destroyed by ficin, papain, bromelin, trypsin  Pk and Luke = globoside collection
and pronase. - At least 2 biosynthetic pathways and genes located at
- Also destroyed by *ZZAP (DTT + ficin/papain) different loci
- Not affected by DTT alone. - Renamed:
 Anti-P -> Anti-P1
 Lectins:  P (+) -> P1
- Anti-M – Iberis amara
 P (-) -> P2
- Anti-N – Vicea graminea, Bauhinia variegate,
 P null -> p
Bauhinia purpurea

S AND s ANTIGENS
- Located on a smaller glycoprotein GPB
- Differentiated by the amino acid at 29th position (S =
methionine; s = threonine)
- Well developed at birth.
- Less easily degraded by enzymes
- Destroyed by ficin, papain, bromelin, pronase and
chymotrypsin.

ANTI-M
- Naturally occurring saline agglutinin
P1 ANTIGEN
- Antibody is most reactive at temperatures below 37C
- Found on fetal RBCs during 12 weeks but weakens
with optimum temperature of 4C and is considered to
with gestational age
be clinically insignificant
- Poorly expressed at birth and fully expressed at 7
- A mixture of IgG and IgM
years
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- Deteriorates rapidly on storage - Antigens are present on all blood cells and have a
- P1 – Lumbricoides terrestris and Ascaris suum wise tissue distribution.
- P1 and Pk – found on Echinococcus granulosus - i antigen is predominantly seen in fetal RBCs which
th
hydatid cyst decreases during the first 18 months of life
- P1 -like antigen – RBc, plasma, pigeon and - I antigen increases until adult proportions are reached
turtledove droppings, egg white of turtledove. - Soluble I antigen is found in milk, saliva, and amniotic
fluid, and a small amount is in plasma.
ANTI-P1  Pathologic Anti-i can cause autoagglutination,
- IgM, naturally occurring in P1(-) individuals vascular occlusion or hemolytic anemia.
- Weak, cold-reactive saline agglutinin (4C) Production may be stimulated by Mycoplasma
- Some are reactive at RT and binds complement at pneumoniae infection (I- like antigen)
37C (detected in AHG phase)  Anti-i - Rare; produce strong reaction with cord
- Associated with Fascioliasis, Clonorchis sinensis RBCs and adult I RBCs; weaker reactions with
and Opistorchis viverinii infection adult I RBCs. IgM, reacts best at 4C, saline
- Neutralized by P1 soluble substances suspended cells. It is associated with infectious
mononucleosis, myeloid leukemia, and alcoholic
ANTI-PP1PK cirrhosis.
- Originally called anti-Tja
- First described on serum of Mrs. Jay (p phenotype KELL BLOOD GROUP SYSTEM
with adenocarcinoma of stomach) - The Kell blood group system is complex and contains
- Tumors contained P system antigens and the many antigens that are highly immunogenic.
antibody has cytotoxic properties that may have - These antigens are the third most potent, after those
helped prevent metastatic growth post-surgery of the ABO and Rh blood groups at triggering an
- Produced by all p individuals w/o exposure immune reaction.
- Reacts with all RBCs except from p individuals - In rare cases, it can cause Hemolytic disease of the
- Components are separable through adsorption • newborn (HDN).
Have IgM and IgG component - HDN cause by Kell immunization tends to result in
- Have wide thermal range and binds complement severe fetal anemia (Maternal anti-Kell taget fetal
(potent hemolysin) RBC precursors, suppressing the fetal production of
- Can cause HTR and HDN RBCs).
- Associated with spontaneous abortion in early - Can cause severe hemolytic transfusion reaction.
pregnancy
K AND k ANTIGENS
ALLOANTI-P  K (Kell)
- Naturally occurring alloantibody in sera of all Pk - Low incidence antigen
individuals. - Second to D to immunogenicity
- Reactivity is similar to anti-PP1Pk (potent hemolysin). - Seen in fetal RBCs in 10 wks
- Reacts with all RBCs except auto control and p - Well developed at birth
phenotype.  k (Cellano)
- High incidence antigen (more common)
AUTOANTI-P - Seen in fetal RBCs in 7 wks
- Associated with PCH - Well developed at birth
- Cold-reactive antibody (IgG with biphasic property)
- Binds RBCs in the cold (4C) and causes hemolysis ANTI-K
(complement activation) at 37C - Most common antibody seen in the blood bank
- Demonstrated by Donath-Landsteiner test - 25 antigens.
- IgG antibody is produced against Kell antigens, IgM is
LUKE (LKE) ANTIGEN AND ANTIBODY uncommon.
- 1965, Tippet described an antibody in serum of - Does not bind complement. If hemolysis does occur, it
Hodgkin Lymphoma patient is extravascular in nature.
- Luke (+), Luke (-), Luke (w) - Associated with Mycobacteria, Enterococcus faecalis,
Morganella morgani, Campylobacter jejuni, and
I BLOOD GROUP SYSTEM Campylobacter coli
- Also known as Ii Blood Group System
- I and i antigens are produced by allelic pairs, they are NULL PHENOTYPE
reciprocal. - The Kell system has a rare null phenotype, Ko, in
- I antigen is formed by the action of an enzyme which RBCs lack all Kell antigens.
(glycosyltransferase), which adds branches onto the - Individuals with this phenotype are healthy but
i antigen. produce anti-Ku when they encounter RBCs that do
express Kell antigens.

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 - Anti-Ku is capable of causing a mild to severe - Epitopes of Fy6 antigen are required for invasion of
transfusion reaction with at least one fatal case being [Link]
reported (5). Therefore, if Ko individuals ever require a - Only murine monoclonal antibodies have defined this
blood transfusion, they should only be transfused with antigen, not human antibodies.
Ko blood products.
DUFFY ANTIBODY
MCLEOD PHENOTYPE AND SYNDROME - Mainly IgG type, and IgM type are rare
- In the RBC membrane, the Kell glycoprotein is - Do not bind to complement
covalently linked to the XK protein, a multipass - Some anti-Fy antibodies have been reported to bind
membrane protein thought to have a role in transport. with complement
- In the absence of XK, a condition called McLeod - Naturally occurring and re acquired by exposure to
syndrome, Kell antigens are only weakly expressed blood transfusion, pregnancy, and transplantation.
a
and the RBCs are abnormal with spiky projections - Fy antibody is most common among other Duffy
(acanthocytosis). antibodies
- Systemic findings include muscular dystrophy, - Majority are reactive with body temperature hence
cardiomyopathy, psychiatric disturbances, and they may cause red hemolysis and are clinically
neurological defects, such as loss of reflexes and significant which may lead to acute and delayed types
movement disorders. of HTR and HDFN

DUFFY BLOOD GROUP SYSTEM KIDD BLOOD GROUP SYSTEM

- Found on the surface of RBCs, vasculature - The Kidd (JK) glycoprotein is the RBC Urea
endothelial cells, alveolar epithelial cells, collecting transporter. Situated in the membrane of RBC, it
tubules of the kidney, and on the surface of Purkinje rapidly transports urea into and out of RBCs,
cells in the brain. maintaining the osmotic stability and shape of the
- Mapped on Chromosome 1 RBC in the process.
- Act as receptors for the chemokines and attract - Also expressed in kidney. It helps kidney to build up
immune system cells. high urea concentration that is needed to produce
- Also acts as receptors for Plasmodium spp. (infection concentrated urine.
resistance from Plasmodium knowlsei and - Although with the above mentioned, those who do not
Plasmodium vivax. have Kidd glycoprotein are healthy and with normal
- May cause Transfusion Reactions (TR) and Hemolytic shape RBC, also normal lifespan. Although they tend
Disease of the Fetus and Newborn (HDFN). not be able to maximally concentrate urine.

DUFFY ANTIGENS KIDD ANTIGENS

a b
 Fya Antigen - Has 3 types of antigen; Jk , Jk and Jk3.
- Expressed on the cord blood RBCs - Well developed in fetal RBCs
- Antigen can be demonstrated on the fetal RBCs as - Not very immunogenic; not destroyed by papain, ficin,
early as 6 weeks gestation. AET, DTT, chloroquine-3-phisphate, glycine-acid
- Adult level of Fya expressions is attained EDTA
approximately 12 weeks after birth.
- Sensitive with ficin, papain, α-chymotrypsin and KIDD ANTIBODIES
a b
resistant to trypsin. - Anti-Jk , Anti-Jk and Anti-Jk3
 Fyb Antigen - Both IgG and IgM, but more common in IgG
- Expressed on the cord blood RBCs - Capable of hemolysis
- Sensitive with ficin, papain, α-chymotrypsin and - Can bind Complement
resistant to trypsin - Common cause of Hemolytic Transfusion Reaction
 Fy3 Antigen (HTR)
- Expressed on the cord RBCs and increases with age - Can be difficult to detect in routine blood cross-
- Resistant with ficin, papain, α-chymotrypsin and matching
resistant to trypsin. - Common cause of delayed HTR
 Fy5 Antigen - Anti-JK3 is rare but can cause immediate and delayed
- Expressed on the cord blood RBCs HTR
- Weak antigenic expression on D negative RBCs and - Rarely cause Hemolytic Disease of the Newborn
absence in Rh null phenotype red cells show possible (HDN) but most cases are in mild in nature.
interaction between Duffy and Rh proteins.
- Resistant to ficin and papain. LUTHERAN BLOOD GROUP SYSTEM
 Fy6 Antigen -
a
Lu is an antigen of the Lutheran blood group system.
- Expressed on the cord blood RBCs It is one of 20 Lutheran antigens on two conserved
- Sensitive with ficin, papain, α-chymotrypsin and red blood cell surface glycoproteins known as
resistant to trypsin. Lutheran (Lu) and basal cell adhesion molecule
(BCAM).
3|Page
 a
- Anti-Lu is rarely clinically significant
a
- Patients with anti-Lu should receive RBC units
crossmatch compatible by IAT by 37C for transfusion
a
- Patients with sickle cell disease who have anti-Lu
a
should be provided with Lu -negative red blood cell
units for transfusion.

a
Anti-Lu
- Naturally occurring IgM
- Some may bind to complement (no hemolysis
reported)
- Often undetected in routine tests
- Resistant to ficin and papain, but sensitive to trypsin,
chymotrypsin, pronase, AET, and DTT

b
Anti-Lu
- Mostly IgG, reactive at 37C and AHG phase
- Restant to ficin and papain, but sensitive to AET and
DTT
- Shortened survival of transfused cells and post-
transfusion jaundice
- Associated with mild Hemolytic Disease of the
Newborn

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 OUTLINE
I. Donor Selections and Screening
 Donor Identification
 Types of Deferral
 Medication Deferral List Other Deferrals:
II. Physical Examination
III. Types of Blood Donation
Avoid intake of aspirin 3 days 2 weeks → if donor received
IV. Donor Reactions prior to donation (not suitable live attenuated or bacterial
for platelet pheresis) vaccine (e.g., rubeola,
DONOR SELECTION AND SCREENING mumps, oral polio, typhoid, or
DONOR IDENTIFICATION yellow fever
- Donor identification must be confirmed by Blood
banks and must be linked to existing donor record 6 weeks deferred for females 4 weeks → infrequent plasma
following pregnancy apheresis
prior the procedure
termination

 List of information needed: 12-month deferral if the 16 weeks → for double unit of
1. Name donor received blood red cells.
2. Age and Birthdate transfusion during her
3. Sex pregnancy
4. Contact Number
8 weeks (56 days) → time 48 hours → time interval
5. Address
interval between allogeneic between apheresis and WB
6. Date/time of collection WB donations. donation
7. Consent form to donate (with signature)
4 weeks → live attenuated 14 – 21 days → smallpox
TYPES OF DEFERRAL vaccine for rubella or vaccine or until the scab has
chickenpox. fallen off
Types Description
- Prospective donor is unable to 12 months deferral → tattoo, 12 months deferral → after
Temporary Deferral donate blood for a definite period ear or body piercing, sexual sexual contact or living with
of time.
- Example: Donor has received a contact with persons w/ person who has hepatitis B,
blood transfusion; defer for 12 HIV/AIDS, treated for syphilis symptomatic hepatitis C or
months from the date of or gonorrhea other hepatitis viruses.
transfusion
- Prospective donor is unable to
Indefinite Deferral donate blood for someone else for 12 months deferral → from 3 years deferral → after
an unspecified period of time due the last day of incarceration completion of treatment and
to current regulatory requirements. (been in juvenile, detention) symptom free if ever had
- Prospective donor will never be
Permanent Deferral eligible to donate blood for
malaria
someone else (may be eligible for
autologous blood donation).
PHYSICAL EXAMINATION OF DONOR

MEDICATION DEFERRAL LIST

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 FORMULAS TO REMEMBER  Apheresis Donation
- Uses an automated cell separator device whose
centrifugal force separates blood into its components
based on differences in densities.
- Methods: intermittent flow and continuous flow
- Plasmapheresis, platelet pheresis, leukapheresis,
double RBC pheresis, stem cell pheresis.
TYPES OF BLOOD DONATION
 Plateletpheresis
- A standard procedure by which platelets are
 Allogeneic Donation
separated from whole blood, concentrated, and
- Blood donation where the blood collected to a person collected.
is given/transfused to another human being. - At least 2 days interval, not more than 2x/week or
- Tests are routinely done to check compatibility of 24x/year.
blood donated before being transfused. - Donors who have ingested aspirin/aspirin-containing
drugs are deferred.
 Autologous Donation - Platelet count is required on first donation if interval is
less than 4 weeks (>150,000/uL).
- Blood donations that a person give for their own use. - Maximum plasma with platelets that can be removed
- Usually done prior surgery, patient’s blood is collected is 500 mL.
and will be used if needed during surgery or after. - If contaminating RBCs are more than 2.0mL, a pilot
- This is to avoid the risks associated with receiving tube must be attached to the product for cross-
blood from other donor. matching purposes.
- This are done for those with rare blood phenotypes - First product to be collected by apheresis.
and those who has multiple antibodies  Occasional donors → not more often than once
every 4 weeks.
 Serial donors → more frequent than once every 4
*Preoperative Autologous Collection- is the weeks.
specific name called for those who will use their own - At least 48 hours interval, maximum of 2
blood for themselves when they are undergoing donations/week
surgery. Usually begins 5-6 weeks before the - Donor plasma must be tested for TP and SPE or
scheduled procedure. quantitative immunoglobulins.

 Acute Normovolemic Hemodilution  Leukapheresis

- More in blood conservation technique than a blood - The only effective way of collecting granuocyte.
- Often used to treat conditions with abnormally high
donation type.
white blood cell counts or to collect white blood cells
- It is when blood of the patient is removed prior the for research or treatments like CAR-T cell therapy.
surgery, and replaced with fluids to maintain the - Therapeutic dose of granulocyte = >1x1011
normal volume of blood circulating. The blood granulocytes per day for 5 days.
removed then returned to the patient after surgery to - Prior to collection, the donor is given:
reduce the need for allogeneic blood transfusion  Hydroxyethyl starch (HES) – sedimenting agent;
enhances separation of WBCs from RBCs during
centrifugation
 Intraoperative Collection
 Prednisone or Dexamethasone – corticosteroids;
- Involves collecting and reinfusing blood lost by a pull granulocytes from the marginal pool into the
patient during surgery. general circulation.
- Blood may be stored at room temperature for 6 hours  Recombinant hematopoietic growth factors –
or at 1C-6C for up to 25 hours provided that latter produce 4-8x the volume of cells in each collection
temperature is begun within 4 hours from end of compared to other agents.
collection.
 Double Red Cells by Apheresis
 Post-Operative Collection
- Two allogeneic/autologous RBC units every 16 weeks
- Blood is collected from a drainage tube placed at the
by automated methods.
surgical site.
- Male donors: at least 5’1” and 130 lbs; female donors:
- Blood is reinfused with or without processing.
at least 5’5” and 150 lbs.
- NO more than 1,400mL be reinfused.
- Hematocrit level (both sexes): at least 40%
- Must be reinfused within 6 hours.
- If collection was discontinued and the amount of RBC
collected was:
 Directed Donation
 <200mL → donor may donate again within 8 weeks
- Same requirements as with allogenic donation except
that the unit is directed to a specific recipient.  >200mL but <300mL → deferred for 8 weeks
- If the donor is a blood relative or HLA-match  >300mL → deferred for 16 weeks
(regardless of immune status), the blood unit must be
irradiated to prevent TA-GVHD.

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 DONOR REACTIONS
 Mild to moderate reactions: includes
syncope/fainting, nausea or vomiting,
hyperventilation, twitching, and muscle spasm.
 Severe reactions: convulsions – caused by cerebral
ischemia, marked hyperventilation, or epilepsy.
 Hematoma – one of most common reaction after
blood collection. Localized collection of blood under
the skin, resulting in a bluish discoloration.

DONOR UNIT PROCESSING

Parameter Remarks
HBsAg - If positive on EIA/CMIA, test must be
repeated in duplicate and a confirmatory
test must be performed.
- If positive on confirmatory, donor is
deferred permanently.
Anti-HBc - Presence in the donor serum suggests
the possibility of HBV infection (acute or
chronic)
- (+) anti-HBc; (+) HBsAg → permanent
deferral
- (+) anti-HBc; (-) HBsAg → not a cause
for donor deferral unless it occurs on
more than one occasion or two
consecutive tests
Anti-HCV and - HCV was initially referred as “non-A
HCV NAT non-B hepatitis”
- NAT for HCV RNA – detects small
amount of viral nucleic acids before
antibodies or viral antigens are
detectable by current methods.
- Recombinant immunoblot assay (RIBA)
– confirmatory test (positive → donor us
permanently deferred)
Anti-HIV 1/2 - If positive for screening, repeat test in
and NAT duplicate (all units must be discarded if
one of the repeat tests is reactive.
- Western blot and IFA – confirmatory
tests.
Anti-HTLV I/II - Causative agent of adult T cell leukemia
and has been associated with HTLV-
associated myelopathy (type 1)
Syphilis - RPR and VDRL → based on the
detection of regain (anti-cardiolipin
antibody).
- FTA-Abs → confirmatory tests; indirect
IFA method
- Treponema pallidum cannot survive in
citrated blood stored in 1-6C for more
than 72 hours
Platelet - Culture method – for apheresis-derived
Bacterial platelets.
Detection - PGD test (verax) – platelets pooled
within 4hrs of transfusion
- Culture method – stored pre-pooled
platelet product
Malaria - HRP II → plasmodium falciparum
trophozoites only • pLDH → P.
falciparum, P. vivac, P. ovale,
[Link]

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 - Additive solutions (AS-1, AS-3, AS-5), a mixture of
glucose, adenine and normal saline provide nutrients
to stabilize cell membrane and maintain the level of
2,3-DPG (minimize “storage lesions”).
- AS-1 and AS-5 contains mannitol as a membrane
stabilizer.
OUTLINE - AS-3 contains citrate and phosphate as membrane
stabilizer.
I. Donor Blood components
II. Whole blood MODIFIED WHOLE BLOOD
III. Packed Red Blood Cells - Whole blood void of cryoprecipitated Antihemophilic
IV. Aliquoted Red Blood Cells Factor (AHF)
V. Leukoreduced Red Blood Cells - Hct = approx. 38%
VI. Washed Red Blood Cells - Stored at 1-6C; stored for 21 days (ACD and CPD)
VII. Frozen Deglycerolized RBCs for 35 days (CPDA-1).
VIII. Platelet Concentrate
IX. Platelet Aliquots IRRADIATED WHOLE BLOOD
X. Leukoreduced Platelet - To inhibit T-cell proliferation.
XI. Granulocyte Pheresis - Expiry date: 28 days from date of irradiation or the
XII. Fresh Frozen Plasma (FFP) original outdate of the unit whichever is sooner
XIII. Liquid Plasma - Minimum dose of gamma radiation = 25 Gy to central
XIV. Cryoprecipitated AHF portion of unit with no less than 15 Gy to any part of
XV. S/D-Pooled Plasma the unit
XVI. FVIII Concentrate
XVII. FIX Concentrate PACKED RED BLOOD CELLS
XVIII. Immune Serum Globulin - For patients requiring an increase in RBC mass.
XIX. Normal Serum Albumin - 200-250 mL of plasma may be removed (CPDA-1)
XX. Plasma Protein Fraction - Additional 50 mL may be removed if additive
solutions are employed (e.g., AS-1)
DONOR BLOOD COMPONENTS - No specific level of Hgb to indicate need for
transfusion.
- 7-8g/dL = patients with heart, lung, or cerebrovascular
disease.
- 7g/dL = for leukemic or surgical patients • <6 g/dL =
critical; for patients w/o disease
- Contraindicated for patients who are well
compensated for anemia.
- 1 unit of PRBC = increase Hb level by 1-1.5 g/dL and
Hct by 3-5% (typical 70kg adult)

ALIQUOTED RED BLOOD CELLS

- For neonates or infants younger than 4 months
- 10-25mL
- CPDA-1
- Transfusion of 10mL/kg with a unit with 80% Hct
(inc in Hb by 3g/dL)
- Expiration date 24 hours and should be stored at 1-
6C until issued.

LEUKOREDUCED RED BLOOD CELLS

WHOLE BLOOD - RBC units with absolute WBC count of <5x10⁶ and
- For replacement of RBC mass and plasma volume. must contain at least 85% of the original RBC mass.
- Dilute into 1:8 (AC-to-blood). - Uses 3rd generation leukocyte reduction filters
- Glucose, adenine, phosphate (additives for RBC (<1x106;1x104)
metabolism during storage). - Pre- or post-storage leukoreduction.
- 1 unit WB = increase Hct by 3-5% or Hb by 1-1.5 g/dL - For the prevention of febrile non-hemolytic transfusion
(for typical 70kg adult) reactions, TA-GVHD, HLA alloimmunization,
- Pediatric patients = 8mL/kg will increase Hb by 1g/dL transmission of CMV, EBV, HIV or HTLV.
and Hct by 3-4%. - 5 x 10⁶ – polyester or cellulose acetate filters
- 5 x 10⁸ – centrifugation

WASHED RED BLOOD CELLS

- For patients with severe anaphylactic transfusion
reactions – for patients with IgA deficiency.
- Removes plasma proteins (e.g., IgA)
- Expiry = 24hours

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 FROZEN DEGLYCEROLIZED RED BLOOD CELLS  Septicemia or bacterial infection
- Stored for up to 10 years unresponsive to antibiotics
- For autologous use  Reversible bone marrow hypoplasia
- Free of WBCs and platelets due to washing
 Reasonable chance of survival
- Cryoprotective agents: penetrating and non-
penetrating
 Penetrating agents (e.g., Glycerol) – small FRESH FROZEN PLASMA (FFP)
molecules that cross the RBC membrane into - For multiple coagulation factor deficiencies in patients
cytoplasm (prevents water from moving out). with liver failure, DIC, vit. K deficiency, warfarin
 Non-penetrating agents (e.g., HES and overdose
dimethylsulfoxide) – large molecules - Given if patients is actively bleeding or time is not
- available for warfarin reversal before surgery.
HIGH GLYCEROL (40% W/V) - Frozen with 8 hours of collection if anticoagulant is
CPD or ACD, stored at 18C for 1 year or -65C for 7
- More widely used
years.
- Increase cryoprotective property of glycerol (allowing
- Thawed at 30-37C
a slow, uncontrolled freezing process)
- Once thawed, unit should be stored at 1-6C for no
- Mechanical freezer (-80C)
more than 24 hours.
- RBCs are frozen within 6 days of collection (CPD,
- 1 unit of FFP = 150-250 mL of plasma approx.
CPDA-1) and up to 42 days (AS-1, AS-3, AS-5).
400mg of fibrinogen, 1 unit of activity/mL of clotting
- Thawing: approx. 30 min (immersion to 37C water
factors.
bath)
- PF24 = frozen within 8 – 24 hours at -18C, contains
- Washing RBCs with solutions of decreasing molarity
all stable proteins found in FFP.
(12% NaCl, 1.6% NaCl, 0.9% NaCl + 0.2% dextrose)
- One deglycerolized, until will expire within 24hours
and is stored at 1-6C. LIQUID PLASMA
- Plasma from whole blood units that can be stored for
LOW GLYCEROL (20% W/V) up tp 5 days after the expiry date of the latter.
- FFP stored for up to 5 days after thawing
- Cryoprotection of glycerol is minimal.
- Keep refrigerated at 1-6C.
- Rapid and controlled freezing is required
- Used as volume expanders or for manufacturing
- Liquid nitrogen (N2)
plasma fractionation products.
- Units are stored at -120C within 6 hours post
collection unless rejuvenated.
- Rejuvenation increases level of 2,3-DPG and ATP in CRYOPRECIPITATED ANTIHEMOPHILIC FACTOR (AHF)
RBCs stored in CPD or CPDA-1 - Cold-precipitated concentrates of FVIII.
- RBCs can be rejuvenated up to 3 days after - Prepared from slow thawing of FFP between 1-6C
expiration and then glycerolized and freeze - Collected in CPDA-1/CPD, suspended in less than
15mL of plasma
PLATELET CONCENTRATE - 80 units of AHF, at least 150mg of fibrinogen.
- 12 months in frozen state
- For actively bleeding thrombocytopenic patients,
- Should be transfused within 6 hours of thawing or 4
cancer patients during radiation and chemotherapy,
hours of pooling
thrombocytopenic preoperative patients.
- Thawed quickly at 37C, stored at 22-24 until
- Not indicated for patients with ITP and DIC.
transfused
- Random donor PC = at least 5.5 x 1010 platelets,
stored at 20-24C with continuous agitation pH > 6.2,
shelf life of 5 days. S/D-POOLED PLASMA
- Plateletpheresis: Single donor PC = 3.0 x 1011 - Consists of pools of no more than 2500 units of ABO-
platelets, for patients who are unresponsive to type specific plasma treated with solvent/detergent
random donor PC due to alloimmunization. in the thawing process.
- To inactivate lipid-enveloped viruses
PLATELET ALIQUOTS - Solvent = tri-n-butyl phosphate; Detergent = Triton x-
100
- For neonates with platelet count <50,000/uL who are
experiencing bleeding
- Expiration is 4 hours from the time the unit was FACTOR VIII CONCENTRATE
spiked. - Indicated for hemophilia A patients
- From large portions of pooled plasma or recombinant
LEUKOREDUCED PLATELETS FVIII.
- For the prevention of febrile non-hemolytic reactions;
< 8.3 x 105 leukocytes FACTOR IX CONCENTRATE
- Pooled platelets and single donor platelets <5x106 - 3 forms: prothrombin complex concentrate, Factor IX
leukocytes concentrate, recombinant Factor IX concentrate
- For those patients with hemophilia B
GRANULOCYTE PHERESIS - PT complex → contains significant levels of vit. K-
dependent factors; from adsorption of pooled plasma
- For patients on chemotherapy for leukemia and bone
with barium sulfate or aluminum hydroxide;
marrow transplant
lyophilized.
- Criteria:
 Fever
 Segmenters count <500/uL
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- FIX concentrate → produced from monoclonal
antibody purification; contains approx. 20-30% FIX;
lyophilized.
- Recombinant FIX concentrate → produced in a
Chinese hamster ovary cell line

IMMUNE GLOBULIN
- Concentration of plasma gamma globulin in aqueous
solution
- Prepared from pooled plasma by cold ethanol
fractionation
- Indicated for [patients with immunodeficiency
diseases and for passive immunization.

NORMAL SERUM ALBUMIN

- Prepared from salvage plasma, pooled, and
fractionated by a cold alcohol process.
- Indicated for patients who are hypovolemic and
hypoproteinemic.

PLASMA PROTEIN FACTOR

- Similar with NSA with fewer purification steps
- 83% albumin, 17% globulin

3|Page
 CAUSES OF SEROLOGICAL CROSSMATCH
- Incorrect ABO grouping of the donor and recipient
- Recipient’s alloantibody reacts with donor RBCs
- Prior coating of donor RBCs with antibodies
- Abnormalities in recipient’s serum
- Contaminants in the test system
OUTLINE
I. Compatibility Testing ANTIBODY SCREENING
II. Serological Screening - Recipient’s serum or plasma
III. Antibody Screening - To detect as many clinically significant unexpected
antibodies as possible
IV. Compatibility Testing in Emergencies
- Clinically significant antibodies = reactive at 37C
V. Compatibility Testing on Plasma Products and/or AHG, known to have caused a transfusion
reaction or unacceptable short survival RBCs
COMPATIBILITY TESTING
- These are tests that are done prior the blood COMPATIBILITY TESTING IN EMERGENCIES
transfusion to ensure safe transfusion and prevent - Sometimes, recipients may require transfusion of
unwanted reactions which may lead to serious, even RBC components prior to completion of pre-
life-threatening reactions. transfusion testing.
- Some lab employs shortened incubation time and
enhancement medium to accelerate Ag-Ab reaction.
RECIPIENT IDENTIFICATION
- ABO and Rh type of recipient must be determined •
- Clerical error → major cause of transfusion In extreme emergencies, group O Rh-negative
associated fatalities (greatest threat). PRBC may be used
- Common cause of errors - Resorting to Rh-positive is best made immediately if
 Misidentification of recipient patient is a man or a woman beyond child-bearing
 Mix-up of samples during processing age.
 Misidentification of recipient when - Administration of RhIg to prevent Anti-D formation
transfusion is given after crisis has been resolved.

RECIPIENT SAMPLE COLLECTION COMPATIBILITY TESTING ON PLASMA PRODUCTS

- Serum or plasma - Not required
- Avoid hemolysis - But If you require plasma you should only receive
- Discrepancies must be resolved before the sample is plasma that doesn't contain an antibody which could
accepted attack the antigens on your own red cells. Group A
- Unlabeled specimen = repeat collection patients have A antigen on their red cells, so they
can't receive group O or group B plasma as the anti-A
DONOR SAMPLE COLLECTION will attack their red cells.
- Must be collected at the same time as the full donor - Donor plasma + recipient RBCs
unit. - To detect ABO incompatibility (should be ABO/Rh
- RBCs for donor pre-transfusion testing can be matched)
prepared from segmented tubing through which donor
blood was collected.
- Donor and recipient sample must be stored for a
minimum of 7 days post transfusion (stoppered,
labeled, 1-6C)

SEROLOGICAL SCREENING
- DONOR = ABO, Rh, tests for blood-borne diseases
- RECIPEINT = ABO, Rh, unexpected antibodies
- Major Crossmatch: patient serum + donor RBCs
(PSDR)
- IS (IgM), 37C (IgG), AHG (non-agglutinating)
- Immediate Spin (IS) Phase
 PSDR + quick centrifugation
 Absence of agglutination or hemolysis
indicates compatibility
 False reactions may be seen in other IS-
reactive antibodies
- Antihuman Globulin (AHG) Phase
 PSDR + IS + 37C + AHG
 Enhancement media may be applied
 Absence of agglutination or hemolysis
indicates compatibility

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 - 2-5% in preservative diluent
- Packaged in sets of 2-3 cells with varied antigen
expression.
- Accompanied by antigen profile sheet
ENHAMCEMENT MEDIA

OUTLINE
I. Antibody Screening
II. Enhancement Media
III. AHG Reagents
IV. Guide to Interpretation
V. Antibody Identification
VI. Evaluation of Panel Reactions
VII. DAT and Elution Techniques ANTIHUMAN GLOBULIN (AHG) REAGENTS

ANTIBODY SCREENING
- It consists of testing blood to check for antibodies
which could cause problems with blood transfusions
or in pregnant women.
- This can be used for investigating potential hemolytic GUIDE TO INTERPRETATION
transfusion reactions, hemolytic disease of the
newborn (HDN), and immune hemolytic anemia

TUBE TECHNIQUE
- Recipient serum + known RBCs
- Enhancement media may be used.
- Hemolysis or agglutination in any phase
- IS + 37C + AHG
 Immediate Spin (IS) Phase
- This is the initial phase where the serum/plasma
and RBCs are mixed and centrifuged
immediately, allowing for the detection of
antibodies that react strongly in room
temperature, often IgM antibodies.
 37C Phase - Agglutination / Hemolysis
- The mixture is then incubated in 37C for a period  Negative (0)
of time allowing for the detection of antibodies  Positive (+1 to +4)
that react optimally in body temperature, often - What Phase did the reaction occur?
IgG antibodies. - Auto control (AC):
 Antihuman Globulin (AHG) Phase  (+) Ab screen; (-) Autocontrol = Alloantibody
- Also known as Coombs test  (+) Ab screen; (+) Autocontrol = Auto- and/or
- The third phase, the sample is then washed with Alloantibody
saline to remove the unbound antibodies and
then the Antihuman Globulin (AHG) reagent is
added. AHG detects antibodies that are bound to
the RBCs but does not cause direct agglutination
(clumping).

*Did more than 1 screening cell react? If so, Did they react at
the same strength?
*Reaction in same phase and same strength = Single
Antibody
*Reaction in more than 1 screening cell (SC) = Multiple
RED BLOOD CELL REAGENTS
Antbodies/Autoantibodies
- Group O, typed for the most common and most
significant RBC antigens.

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Agglutination vs Rouleaux
- Rouleaux formation is a physical phenomenon where
RBCs stack due to increased plasma protein. It is
reversible by adding saline.
- Agglutination, on the other hand, is immune-
mediated process where RBCs clump together due
to antibodies binding to them. It is not reversible by
adding saline.

ANTIBODY IDENTIFICATION
- Reagents: 11-20 Group O cells with varied antigen
expression.
- Pattern of antigen expression should be diverse
enough to distinguish one antibody from another.
- A profile sheet specifying the antigens on each cell
and providing a place to record reactions
accompanies each panel.

EVALUATION OF PANEL REACTIONS

- IS Phase = IgM (clinically significant)
- 37C/AHG Phase = IgG (clinically significant)
- May also indicate antibodies showing dosage effect
- Pattern of reactivity usually matches the other
patterns exactly = SINGLE ALLOANTIBODY

DAT AND ELUTION TECHNIQUES

- Detection of antibodies coating RBCs when
investigating suspected hemolytic transfusion
reactions, HDN, AIHA/DIHA
- Direct AHG Test (DAT) – detects in vivo attachment
of antibodies to RBC surface
- Elution – dissociating attached antibodies from the
cell surface to allow identification.
 Eluate – solution with freed antibodies
 Total elution – antibodies are released, and
the RBC antigens are destroyed
 Partial elution – antibodies are removed but
RBC antigens remain intact

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 - Usually occurs within 1-2 hours , but may still occur within
minutes
- Can be intravascular or extravascular
- A positive Direct Antiglobulin Test (DAT), evidence of
hemolysis (elevated lactate dehydrogenase, elevated
bilirubin, low haptoglobin, hemoglobinemia) and acute renal
OUTLINE
impairment can help diagnosis AHTR.
I. Transfusion Reactions
 Hemolytic Transfusion Reaction (HTR) DELATYED HEMOLYTIC TRANSFUSION REACTION
 Immediate HTR - Most often a result of anamnestic response in a patient who
 Delayed HTR have been previously sensitized by pregnancy, transfusion,
 Febrile Non-hemolytic Transfusion Reaction transplantation, and in whom antibody is not detectable by
 Allergic (Urticaria) Transfusion Reaction standard pre-transfusion methods
 Anaphylactic Transfusion Reaction - 2 phases: primary alloimmunization and
 TRALI secondary/anamnestic response
 TACO - In secondary response, 3-7 days post transfusion are
 Bacterial Contamination necessary for production of antibodies and to cause signs
 Physically/Chemically Induced Transfusion and symptoms of extravascular hemolysis.
Reactions (PCITR) - A positive DAT may be delayed, and the diagnosis can be
 Post-transfusion Purpura difficult
 TA-GVHD
 Iron OverLoad FEBRILE NON-HEMOLYTIC TRANSFUSION REACTION
 - Most commonly encountered type of transfusion
II. Transfusion Reaction Investigations reaction.
- Defined as 1C temperature rise associated with transfusion
TRANSFUSION REACTIONS and having no medical explanation other than blood
- Any adverse effect that occur during or after transfusion of component transfusion.
blood which may be cause by immune or non-immune - Caused by anti-leukocyte antibodies present in the
reaction. patient’s plasma
- Prior alloimmunization is the causative stimulus.
- Febrile mechanism still not fully explained
- Febrile reaction may follow complement activation and
production of IL-1 and prostaglandin.

ALLERGIC (URTICARIA) TRANSFUSION REACTION

- Commonly reported as Febrile Non-Hemolytic Transfusion
Reaction
- One of the 2 most common reactions yet definitive causes
are unknown.
- Two possible etiologies:
 Donor plasma has allergen with IgE/IgG
antibodies in patient’s plasma
HEMOLYTIC TRANSFUSION REACTION  Donor plasma has IgE/IgG that combine with
- This happens when the recipient’s body recognizes the allergens in patient’s plasma
transfused RBCs are foreign and attacks the, leading to
RBCs destruction and breakdown. - Histamine – primary mediator of the allergic response
- Occur during transfusion (immediate) or 3-7 days post-
transfusion (delayed) ANAPHYLACTIC TRANSFUSION REACTION
- Often occur with transfusion of incompatible RBCs - A more severe, potentially life-threatening allergic reaction
- Most common cause is ABO incompatibility after blood transfusion which may occur immediately after
the transfusion.
IMMEDIATE HEMOLYTIC TRANSFUSION REACTION - It is characterized as rapid onset with symptoms like hives,
- Also known as, Acute Hemolytic Transfusion Reaction difficulty of breathing, drop of blood pressure, loss of
(AHTR) consciousness, nausea/vomiting, rapid heart rate, and in
- It is a life threatening reaction which may occur within the severe cases, shock
24 hours after the blood was transfused to the recipient. - Probable causes are :
- Most common cause is ABO incompatibility as well.  IgA deficiency – a recipient with IgA deficiency
- In anesthetized patients → abnormal bleeding at the develops antibodies against the IgA from the
surgical wound, hemoglobinuria, hypotension. donor’s plasma

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  Anhaptoglobinemia – a rare condition, where the - Rapid onset of thrombocytopenia due to production of
recipient lacks haptoglobin (protein that binds to platelet alloantibodies
hemoglobin) that causes anaphylactic shockk - Anti-PLᴬ¹ → most frequently identified platelet alloantibody
 Other Allergic Reaction – other
protein/substance present in the blood transfused TRANSFUSION ASSOCIATED-GRAFT VS HOST DISEASE
( TA-GVHD )
- Anaphylactic – IgA-deficient with anti-IgA - Complication of blood transfusion or BM transplantation
- Anaphylactoid – normal levels of IgA but a limited type- - Death is due to infection or hemorrhage secondary to BM
specific anti-IgA reacts with light chains of the donor IgA. aplasia
- Mediated by histamine and leukotriene - Blood component irradiation
 Best current technology to reduce risk of TA-
TRANSFUSION ASSOCIATED ACUTE LUNG INJURY GVHD
( TRALI )  Inactivates lymphocytes
- A noncardiogenic pulmonary edema (fluid buildup in the  25-35Gy
lungs) and hypoxia (low blood oxygen) that develops during
or shortly after a blood transfusion. IRON OVERLOAD
- Similar to adult respiratory distress syndrome (ARDS) - Long term complication of RBC transfusion
- It is a serious complication but is relatively rare in - Also known as Transfusion Hemosiderosis
occurrence - 225 mg Fe/RBC unit
- Pathophysiology is not well understood
- Consistent finding is leukocyte antibody in donor or patient TRANSFUSION REACTION INVESTIGATION
plasma - Adverse clinical manifestation post transfusion must be
- Associated with anti-WBC antibodies promptly evaluated
- TRI is necessary for:
TRANSFUSION ASSOCIATED CIRCULATORY OVERLOAD  Diagnosis
( TACO )  Selection of appropriate therapy
- This happens when the body can’t handle the extra fluid  Transfusion management
from the transfusion, causing the fluid to build up in the  Prevention of future transfusion reactions
lungs (and other parts of the body). - Specimen required during TRI
- It can lead to respiratory distress, swelling, and increase in  Clotted blood drawn 5-7 hours post transfusion
blood pressure.  First voided post-transfusion urine
- Usually occurs within 6-12 hours after transfusion
 Other specimens collected at various times that
- Commonly occurs in recipients with cardiovascular/kidney
are considered appropriate to TRI.
disease, children, and elderly
- May also be caused by transfusion of blood unit too fast
DIRECT AHG TEST
(>200mL/hr)
- Washed RBCs are indicated to reduce plasma oncotic load

BACTERIAL CONTAMINATION
- A type of specific reaction that can have a rapid onset and
lead to death
- Yersinia enterocolitica → most common cause
- Caused by endotoxin produced by psychrophilic bacteria
(Pseudomonas spp., Escherichia coli, Yersinia
enterocolitica)

PHYSICALLY/CHEMICALLY INDUCED TRANSFUSION

REACTION ( PCITR )
- Caused by broad range of physical or chemical factors that
either affect a blood component or are a consequence of
the transfusion event.
- RBC Damage
 Hypertonic/hypotonic solutions
 Heat damage from blood warmers
 Freezing damage in the absence of cryoprotective
agents

POST TRANSFUSION PURPURA - Done using post-transfusion specimen

- Rare complication of blood transfusion involving platelets - Also known as Direct Coombs test

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- It detects antibodies or complement proteins attached on
the surface of RBCs in vivo
- DAT may yield negative result if incompatible transfused
cells have been immediately destroyed
- MF → agglutinated donor RBCs and unagglutinated patient
RBCs

ABO / RH
- Patient pre/post transfusion specimen + donor segment
sample
- Misidentification of any other patient sample or donor unit
should be ruled out

COMPATIBILITY TESTING
- Patient pre/post transfusion specimen + donor unit involved
- Incompatible crossmatch (pre trans specimen) → original
error
- Incompatible crossmatch (post trans specimen) → possible
anamnestic response

ANTIBODY SCREENING/IDENTIFICATION
- False negative pre-transfusion Ab screen
- Pre/post → POSITIVE → antibody ID is essential to
determine Ab specificity to avoid another reaction when
patient requires further transfusion

URINE TEST
- First voided post-transfusion urine
- Free hemoglobin (to established hemoglobinuria)
- It is crucial to determine if there's hemolysis (breakdown of
red blood cells) and to differentiate between hematuria
(blood in urine due to urinary tract issues) and
hemoglobinuria (hemoglobin in urine due to red blood cell
breakdown)

BILIRUBIN TEST
- Bright/ deep yellow serum → RBC hemolysis
- It is also a part of test for suspected blood transfusion
reaction and also to evaluate and assess for hemolysis
- Elevated bilirubin levels can indicate increased breakdown
of RBCs.

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