Blood Banking Review

Medical Technology Review
by ANDRE KARL S. FACULIN, RMT
1
APRIL 7, 2014
 8:00-9:00am PRE-TEST
 9:00-10:30am LECTURE 1
 10:30-11:00am BREAK
 11:00-12:00pm LECTURE 2
 12:00-1:30pm BREAK
 1:30-3:00pm LECTURE 3
 3:00-3:30pm BREAK
 3:30-5:30pm LECTURE 4
2
APRIL 8, 2014
 8:00-10:30am LECTURE 5
 10:30-11:00am BREAK
 11:00-12:00pm LECTURE 6
 12:00-1:30pm BREAK
 1:30-3:00pm LECTURE 7
 3:00-3:30pm BREAK
 3:30-5:30pm LECTURE 8
3
APRIL 9, 2014
 8:00-10:30am LECTURE 9
 10:30-11:00am BREAK
 11:00-12:00pm LECTURE 10
 12:00-1:30pm BREAK
 1:30-3:00pm LECTURE 11
 3:00-3:30pm BREAK
 3:30-5:00pm LECTURE 12
 5:00-6:00pm POST-TEST
4
20%= Immunology, Serology and Blood
Banking
50%=Immunology & Serology

50%=Blood banking
5
6
1. ABO and Rh blood group systems 5%
2. Other major blood group systems: Kell, Duffy, Kidd, Lewis, MNSs, 3%
Lutheran, P, I
3. Minor blood group systems: Diego, Cartwright, Chido, XG, Scianna, 1%
Gerbisch, Milton, Knops, Bg, Indian, etc.
4. Basic genetics 2%
5. Blood donor selection and processing 5%
6. Blood preservation and banking 5%
7. Component preparation 5%
8. Transfusion therapy 2%
9. Transfusion reactions 3%
10. Transfusion-transmitted diseases 3%
11. BB techniques and procedures: typing, compatibility testing, antibody 8%
detection and identification
12. Hemolytic disease of the newborn and autoimmune hemolytic anemia 4%
13. Quality management (structure, set-up/ equipment) 4%
7
8
 Egyptians bathe blood
 Aristocrats drank it
9
 1492– First recorded blood transfusion
• Pope Innocent VII and 3 young men
 1628– William Harvey discovered

circulation of blood
 1665–
Richard Cower: blood transfusion
between dogs
10
 1667– Jean Baptiste Denys: blood
transfusion between a sheep and a
human
 1818–
James Blundell: blood transfusion
between humans
 1869–
First non-toxic anticoagulant:
sodium phosphate
• Braxton Hicks
11
 1901– Karl Landsteiner
 A, B, O blood types
 Book: ___________________________
 1902: vonDescatello & Sturle/

von Decastello & Sturli
 AB blood type
 Edward Lindemann: first to use

appropriate device (multiple syringe and
cannula)
12
 1907–Ludvig Hektoen: Crossmatching
before transfusion
 1908– Carlos Moreschi: Antiglobulin

reaction
13
 1914– Hustin: use of sodium citrate as Ac
 1915–
Levisohn: determined the
minimum amount for anticoagulation and
demonstrated its non-toxicity
 1916– Rous and Turner: citrate-dextrose

solution
14
 1930’s–
function of glucose in RBC
metabolism was known
 1932– First blood bank in Leningrad,

Russia
 1943–Loutit and Mollison introduced

formula for ACD
15
 1945– Coomb, Mourant, Race: Antihuman
globulin reagent
 1954-- Cryoprecipate
 1957– Gibson introduced improved

citrate phosphate dextrose (CPD)
16
 1960– Plasmapheresis (therapeutic)
 1967– Rh Immunoglobulin
 1979– CPDA-1
17
18
 Studyof inheritance or the transmission
of characteristics from parents to
offpsring
• Population
• Cellular
• Molecular
 Importantin study of antigen inheritance
and inherited disorders
19
20
Nitrogenous
bases
+
Sugar backbone
+
Phosphate
21
22
23
Genes
• Basic unit of inheritance
• Segments of DNA arranged along the
chromosome at a specific position
• Encode certain traits or visible
characterisitcs
24
Genetic loci
• Sites of a gene in a chromosome
25
Alleles
• Alternate forms of a gene that differ in their
nucleotide sequence at a given locus
 Homozygous (identical alleles)

 Heterozygous (nonidentical alleles)
 Dominant
 Recessive
26
Codominance
• Equal expression of two different inherited
alleles
• Common among blood group antigens
27
Incomplete dominance
• Intermediate inheritance in which one allele
for a specific trait is not completely dominant
over the other allele
• E.g. Red flowers + White flowers = Pink
flowers
28
Alleles
• Polymorphic
 Two or more alleles at a given locus
 Expresses two or more phenotypes
 E.g. ABO blood group system
29
Alleles
• Antithetical
 Opposite allele
 Used when referring to antigens produced by
allelic genes
 E.g. Kpa antigen is antithetical to Kpb antigen
30
Amorphic
• Gene that does not express a detectable
product
• Silent gene
• E.g. ‘O’ in ABO blood group system
‘d’ in Rh blood group system
31
Dosage effect
• Presence of homozygous genotype can
express itself with more antigen than
heterozygous genotype
Duffy loves
32
Linked genes
• When two genes are inherited together by
being very close on a chromosome
33
Haplotype
• Linked set of genes inherited together
because of their close proximity on a
chromosome
• E.g. MNS blood group system
 M and N are alleles on one gene
 S and s on another
34
 Cis
• Two or more genes on the same chromosome of
a homologous pair
 Trans
• Inherited on opposite chromosomes of a
homologous pair
35
D and C in cis D and C in trans
36
Linkage disequilibrium
• Phenomenon of antigens occuring at a
different frequency in the population,
depending on whether they were inherited
by linked or unlinked genes
 MNS blood group system
 If unlinked= 17% frequency
 If linked= 24 % frequency
37
Cell Division
• Allows the genetic material in cells to be
replicated
 Mitosis= somatic cells, 2N
 Meiosis= gametes, N
38
Phenotype
• Physical or observable expression of
inherited traits
 Red cells + antisera = presence or absence of
hemagglutination
39
Genotype
• Actual genetic make-up
 Family studies
Figure. Pedigree Chart

40
41
Punnett Square
• Illustrates the probabilities of phenotypes from
known or inferred genotypes
A O
B
O 42
Blood Group System
• Groups of antigens on red cell membrane
that share related serologic properties and
genetic patterns of inheritance
43
 Austrian monk, Gregor Johann Mendel
 Sweet pea plants
 Mendel’s law of inheritance
• Law of independent segregation

• Law of independent assortment
44
Parental RR rr
Gametes
R r
First-filial Rr
R r
Second-filial
R RR Rr
r Rr rr
45
Parental RRYY rryy
Gametes
RY ry
First-filial RY, Ry, rY, ry
RY Ry rY ry
Second-filial RY RRYY RRYy RrYY RrYy
Ry RRYy RRyy RrYy Rryy
rY RrYY RrYy rrYY rrYy
ry RrYy Rryy rrYy rryy
46
Incomplete dominance
Codominance
Multiple alleles
47
Autosomal: 22 pairs of autosomes
(non-sex determining chromosomes)
• Autosomal dominant
 Huntington disease, polycystic kidney disease
• Autosomal recessive
 Cystic fibrosis, Tay-sachs, PKU
48
X-linked: X chromosome
• X-linked dominant
 Xga blood group system,
hypophosphatemic rickets
• X-linked recessive
 Hemophilia, SCID
49
Blood group system Chromosome
Rh, Duffy 1
Gerbich 2
MNS 4
Chido/Rodgers 6
Kell 7
ABO 9
Diego 17
Kidd 18
H, Lewis, Lutheran, 19
Landsteiner-Wiener
P 22
50
 Used to estimate the frequency of genetic
diseases and establish probability tables
for forensic and paternity calculations
(p+q)2= p2 (AA) + 2pq (Aa) + q2 (aa)
where:
p is the frequency of allele A
q is the frequency of allele a
51
 Criteria for use of Hardy-Weinberg formula
• Large population
• Random mating
• No mutation in parents/ offspring
• No migration, differential fertility or mortality of
genotypes
52
53
 International
Society of Blood Transfusion
(ISBT) Terminology for Red Blood Cell
Surface Antigens in Blood Group Systems
54
ISBT System Number System Chromosome
001
002 MNS 4
003 P 22
004
005 Lutheran 19
006 Kell 7
007 Lewis 19
008 Duffy 1
009 Kidd 18
010 Diego 17
011 Cartwright 7
012 Xg Xp
55
001 ABO 9
002 MNS 4
003 P 22
004 Rh 1
005 Lutheran 19
006 Kell 7
007 Lewis 19
008 Duffy 1
009 Kidd 18
010 Diego 17
011 Cartwright 7
012 Xg Xp
56
013 Scianna 1
014 Dombrock 12
015 Colton 7
016 Landsteiner-Wiener 19
017 Chido/Rodgers 6
018 H 19
019 Kx Xp
020 Gerbich 2
021 Cromer 1
022 Knops 1
023 Indian 11
024 Ok 19
57
025 Raph 11
026 John Milton Hagen 15
027 I 6
028 Globoside 3
029 GIL 9
030 Rh-associated
glycoprotein
58
59
 Blood
group antigens/ agglutinogens/
immunogens
• Expressed in the RBC membrane except Lewis
• Autosomal codominant
• Proteins: Rh, MN
• Glycoproteins: HLA
• Glycolipids: ABH, Lewis, Ii, P
60
 Blood group antibodies/ agglutinins
61
Naturally-occurring Immune
Found in the serum of Found in the serum of
individuals who have individuals who have
never been previously been transfused or
exposed to RBC antigens pregnant, not generally
by means of transfusion, found in nature
injection or pregnancy
Mostly IgM cold agglutinins Mostly IgG warm antibodies

Isoagglutinins
62
 Reagent antibodies (antisera)
Polyclonal Monoclonal
Serum antibodies that are Same variable region and has
produced in response to a single epitope specificity
single antigen with more Produced by isolating
than epitope individual B cells from a
polyclonal population and
propagating them in cell
culture with hybridoma
technology
Diversity is not optimal in the Preferred due to high
laboratory specificity and uniform
reactivity
63
64
IgM IgG
• D, C, E, c, e, K,
• A, B, H, Lewis,
Fy, Jk, Ss,
Ii, P1, M, N, Lua
Lewis, Lub
• Immediate
• Antiglobulin
spin
phase
• Room temp
• Body temp
• (22-24°C)
• (37°C)
65
Unexpected antibodies
• All other antibodies directed
against RBC antigens that must be
detected and identified prior to
transfusion
• Highly varied as they may be of
the IgM or IgG class
• Antibody screening
66
Alloantibody Autoantibody
Produced after exposure Produced in response to
to genetically different self antigens
or non-self antigens of
the same species Pan- or Polyagglutinins are
autoAb without
detectable specificity
67
1
Genetics and Immunology
68
69
Most important in transfusion and
transplantation
• Antigens:
 RBC
 Secretions
• Antibodies:
 Histoblood Group Antigens
• Present on all tissues and organs of the body
• May be expressed in secretions depending on
secretor status
70
 ABO Forward Grouping
BLOOD REACTION WITH REACTION WITH REACTION WITH

GROUP ANTI-A ANTI-B ANTI-AB
A
B
AB
Note: If label of anti-sera is removed, check with controls!
71
 ABO Reverse Grouping
BLOOD GROUP REACTION WITH A CELLS REACTION WITH B CELLS
AB
72
73
Blood Group Whites Blacks
O 45% 49%
A 40% 27%
B 11% 19%
AB 4% 4%
74
75
76
Universal donor
Universal recipient
Universal donor for RBCs

Universal recipient for RBCs
Universal donor for plasma/plasma products

Universal recipient for plasma/plasma products
77
Universal donor O
Universal recipient AB
Universal donor for RBCs O

Universal recipient for RBCs AB
Universal donor for plasma/plasma products AB

Universal recipient for plasma/plasma products O
78
79
Grading Interpretation
0 No agglutination or hemolysis
W+ Tiny agglutinates, turbid background
1+ Small agglutinates, turbid background
2+ Medium-sized agglutinates, clear background
3+ Several large agglutinates, clear background
4+ One solid agglutinate
80
 Cards containing microtubes with gel
particles and reagent are added with
serum or cell suspensions, incubated and
centrifuged.
 Controlled centrifugation of RBCs
through a dextran-acrylamide gel
81
82
Grading Interpretation
Mixed-field Layer of red cell agglutinates at the top of the gel column
accompanied by a pellet of unagglutinated cells in the
bottom of the microtube
Negative Red cells forming a well-delineated pellet in the bottom
of the microtube. The gel above the red cell pellet is clear
and free of agglutinates
1+ Red cell agglutinates predominantly observed in the
lower half of the gel column with red cells also in the
bottom
2+ Red cell agglutinates dispersed throughout the gel
column with few agglutinates at the bottom
3+ Predominant amount of agglutinated red cells towards the
top of the gel column with few agglutinates staggered
below the thicker band
4+ Solid band of agglutinated red cells at the top of the gel
column with no red cells usually visible at the bottom
83
ABO
H Blood Group I/i

System:
Common
carbohydrate
structure
Lewis P1
84
N-
Gal Gal Glu RBC
AGlu
85
Type Linkage Found in
Type 1 β 13 linkage Body fluids,
secretions
Type 2 Β 14 linkage RBCs, body
fluids,
secretions
86
Gene Glycosyltransferase Immunodominant Acceptor Antigen
sugar
H α-2-L-fucosyltransferase L-fucose Precursor H
A α-3-N-acetylgalactosaminyltransferase N-acetyl-D-galactosamine H A
B α-3-D-galactosyltransferase D-galactose H B
AB α-3-N-acetylgalactosaminyltransferase N-acetyl-D-galactosamine H A, B
α-3-D-galactosyltransferase D-galactose
O H
87
Gene Glycosyltransferase Immunodominant Acceptor Antigen
sugar
H α-2-L-fucosyltransferase L-fucose Precursor H
A α-3-N-acetylgalactosaminyltransferase N-acetyl-D-galactosamine H A
B α-3-D-galactosyltransferase D-galactose H B
AB α-3-N-acetylgalactosaminyltransferase N-acetyl-D-galactosamine H A, B
α-3-D-galactosyltransferase D-galactose
O H
Chromosome 9: A, B
Chromosome 19: H
88
N-
Gal
AGlu Gal Glu
RBC
α12
Fuc
89
N-
N-
Gal
AGlu Gal Glu
RBC
α12
AGal
Fuc
α13
90
N-
Gal
AGlu Gal Glu
RBC
α12
Fuc
91
N-
Gal
Gal
AGlu Gal Glu
RBC
α12
Fuc
α13
92
N-
Gal
AGlu Gal Glu
RBC
α12
Fuc
93
N-
N-
AGal
Gal
AGlu Gal Glu
RBC
α13
α12
Gal Fuc
94
95
Phenotype Possible Genotypes
A1
A2
A1B
A2B
B
O
96
Phenotype Possible Genotypes
A1 A1A1, A1A2, A1O
A2 A2A2, A2O
A1B A1, B
A2B A2B
B BB, BO
O OO
97
A1 A2
A1 A A 1 A AAAA
A1 A A 1 A1 AAAA
A A1 A A1 AAAA
Blood Group Antigen Present Anti-A Anti-A1 lectin

(Anti-A plus Anti-A1)
A1
A2
98
A1 A2
A1 A A 1 A AAAA
A1 A A 1 A1 AAAA
A A1 A A1 AAAA
Blood Group Antigen Present Anti-A Anti-A1 lectin

(Anti-A plus Anti-A1)
A1 A, A1 + +
A2 A + 0
99
 Proteinspresent in plants (usually seeds),
 Bind specifically to carbohydrate
determinants and agglutinate
erythrocytes through their cell surface of
oligosaccharide determinants
10
0
Lectin Serologic specificity
Ulex europaeus
Vicia graminea
Iberis amara
Dolichos biflorus
Griffonia simplicifolia
Helix pornatia Anti-A, Th, Cad
Arachis hypogaea Anti-T, Tk, Th, Tx
Glycine soja Anti-T, Tn
Leonorus cardiaca Anti-Cad
Salvia sclerea Anti-Tn specific
Salvia horminum Anti-Tn, Cad (separable)
Vicia cretia Anti-T, Th
10
1
O>A2>B>A2B>A1>A1B
10
2
Genotype Secretor state %
Se Se
Se se
se se
10
3
 Specimen: Saliva
 Principle: Hemagglutination Inhibition
(+)______________
(–) ______________
10
4
1. Collect 2 to 3 mL saliva in a test tube
2. Centrifuge at 900 to 1000xg for 8 to 10
minutes
3. Transfer supernatant and place
stoppered tube in a boiling water bath
for 10 minutes to inactivate enzymes
4. Recentrifuge and collect clear
supernatant
5. Dilute saliva with NSS
10
5
6. Add one drop of diluted antiserum.
7. Add one drop of supernatant saliva. Mix
and incubate for 8 to 10 minutes.
8. Add one drop of appropriate indicator
cells
9. Mix and incubate at RT for 30-60 mins
10. Centrifuge
11. Observe for macroscopic agglutination
10
6
ABO Group Secretor A cells B cells O cells
A Se Se
B se se
AB se se
O Se se 10
7
Anti-A Anti-B Anti-H
ABO Group Secretor A cells B cells O cells

A Se Se No agg (+) Agg (-) No agg (+)
B se se Agg (-) Agg (-) Agg (-)
AB se se Agg (-) Agg (-) Agg (-)
O Se se Agg (-) Agg (-) No agg (+) 10
8
ABO Group Secretors ABH Substances in Saliva
A A, H
B B, H
AB A, B, H
O H
Non-secretor None
10
9
Saliva Tears Urine
Digestive
Bile Milk
juices
Pathologic fluids:
Amniotic pleural, peritoneal,
fluid pericardial, ovarian
cyst
11
0
Weakly
agglutinated
A3, Aend, Ax
Red cell
reactions with
anti-A and anti-B
Adsorbs and
No agglutination
elutes anti-A Am, Ay, Ael
11
1
Mixed-field A3
≤10% red cells show
Weakly
agglutinated
very weak mf
agglutination
Aend
Red cell reactions
with anti-A and
anti-B Weak
agglutination with
anti-A,B only
Ax
No agglutination
11
2
Easily adsorbed
Weakly
agglutinated
and eluted;
Secretors Am
Red cell reactions demonstrate A subs
with anti-A and
anti-B
Secretors
No agglutination demonstrate small
amount of A subs
Ay
Secretors contain
only H subs Ael
11
3
Weakly
agglutinated
B3 , Bx
Red cell
reactions with
anti-A and anti-B
Adsorbs and
No agglutination
elutes anti-A Bm, Bel
11
4
Mixed-field B3
Weakly
agglutinated
Red cell reactions Weak
with anti-A and
anti-B
agglutination with
anti-A,B only
Bx
No agglutination
11
5
Weakly
agglutinated
Easily adsorbed
Red cell reactions
with anti-A and
anti-B
and eluted;
Secretors Bm
demonstrate A subs
No agglutination
Secretors contain
only H subs Bel
11
6
 First
reported by Bhende in 1952 in
Bombay, India
 hh genotype or H null
 No H antigens formed
• Lacks H, A and B antigens
 Anti-A, anti-B, anti-AB, anti-H
present
 Blood type specific during transfusion
(Oh to Oh)
 Mistyped as Group O
11
7
Anti-A Anti-B Anti-H A cells B cells
Group O - - + +
Oh - - + +
11
8
Anti-A Anti-B Anti-H A cells B cells
Group O - - + + +
Oh - - - + +
11
9
 GroupI Discrepancies: WEAKLY REACTING
OR MISSING ANTIBODIES
• Newborns
• Elderly patients
• Patients with leukemia demonstrating
hypogammaglobulinemia
• Patients with lymphoma demonstrating
• Patients using immunosuppressive drugs that yield
• Patients with congenital hypogammaglobulinemia
• Patients with immunodeficiency diseases
• Patients with bone marrow transplantation
12
0
 Group O newborn or elderly
Anti-A Anti-B A1 cells B cells O cells Autocontrol
0 0 0 0 0 0
0 3+ 0 0 0 0
12
1
 Chimera Twins
Anti-A Anti-B Anti-AB A1 cells B cells RBC %
0 2+mf 2+mf 4+ 0 70% B; 30% O
0 +wk +wk 4+ 0 30% B; 70% O
12
2
 Group
II Discrepancies: WEAKLY
REACTING OR MISSING ANTIGENS
• Subgroups of A and/or B
• Leukemias (weakened A or B antigens)
• Hodgkin’s disease
• Excess amounts of blood group-specific soluble
substances (BGSS) in association with certain
diseases such as carcinoma of the stomach and
pancreas
• Acquired B phenomenon
• Antibodies to low incidence antigens 12
3
 Patient with leukemia
Px phenotype Anti-A Anti-B A1 cells B cells
A +mf 0 0 3+
B 0 +/- 4+ 0
12
4
 Acquired B phenomenon
Anti-A Anti-B A1 cells B cells
4+ 2+ 0 4+
12
5
 Group
III Discrepancies: PROTEIN OR
PLASMA ABNORMALITIES RESULTING
TO ROULEAUX FORMATION
• Elevated levels of globulin from certain disease
states: MM, Waldenstrom’s macroglobulinemia,
other plasma cell dyscrasia, advance Hodgkin’s
lymphoma
• Elevated levels of fibrinogen
• Plasma expanders such as dextran and
polyvinylpyrrolidone
• Wharton’s jelly
12
6
 Plasma abnormalities
Anti-A Anti-B A1 cells B cells
4+ 2+ 2+ 4+
12
7
 Group
IV Discrepancies:
MISCELLANEOUS
• Polyagglutination
• Cold reactive antibodies
• Unexpected ABO isoagglutinins
• Antibodies other than anti-A and anti-B may
react to form antigen-antibody complexes that
may then adsorb into patient’s RBCs
• RBCs with the cis AB phenotype
12
8
 Cold autoantibody
0+ 4+ 4+ 1+ 1+ 1+
12
9
 Group AB with alloantibody
4+ 4+ 2+ 0+ 2+ 0
13
0
 Inadequate identification of blood
specimens, test tubes or slides
 Cell suspension either too heavy or too light
 Clerical errors
 A mix-up in samples
 Missed observation of hemolysis
 Failure to add reagents
 Failure to follow manufacturer’s instructions
 Uncalibrated centrifuge
 Contaminated reagents
 Warming during centrifugation
13
1
Discrepancy Resolution
Group I Check patient’s history
Check for technical discrepancy
Incubate px serum with reagent A1 and B cells at RT and at
4⁰C for 15-30 minutes
Group II Check patient’s history
Incubate test mixture at RT and at 4⁰C for 15-30 minutes
Group III Check patient’s history
Saline dilution or saline replacement technique
For cord cells: wash with saline for 6 to 8 times
Group IV Check patient’s history
Warm technique
DTT treatment
13
2
 Inheritanceof both AB genes from one
parent carried on one chromosome and
an O gene inherited from the other
parent
13
3
Group: AB
Genotype: ABO
13
4
13
5
2 ABO Blood Group System
13
6
13
7
 One of the most complex system because
nearly 50 different Rh antigens have been
identified
 Antigen:
 Antibody:
13
8
 Levine and Stetson
• HTR in an obstetrical patient
• Antibody isolated from the mother
• Postulated that the fetus and father possessed a
common factor that the father lacked
 Landsteiner and Wiener
• Rh
Rabbit  Rhesus monkey blood thought as anti-Rh
immunized
13
9
 Anti-LW
• Originally defined as anti-Rh in early
experiments involving rabbits immunized with
Rhesus Monkey blood
Anti-LW is different from anti-D

 Anti-LW will agglutinate Rh (+) and Rh (-)
cells except Rh null cells
14
0
 Fisher-Race (DCE)
• Based on the theory that antigen of the systems
were produced by three closely linked set of
alleles
• Each gene was responsible for producing a
product or antigen on the red cell surface
D
• D
gene
C/c
• C/c
gene
E/e
• E/e
gene 14
1
 Wiener (Rh-Hr)
• Gene responsible for defining Rh actually
produced an agglutinogen that contained a
series of blood factors, in which each factor is an
antigen recognized by an antibody
Factor Rh0 Rh0
Rh0 gene Factor hr’ hr’
Factor hr’’ Hr’’

14
2
14
3
 Rosenfield
• Number assigned to each antigen of the Rh
system in order of its discovery
• No genetic basis only indicates presence of Rh Ag
 ISBT : Numeric technology

• Adopted a six-digit number for each blood group
specificity
• First three numbers represent system and the
remaining three the antigenic specificity
14
4
Fisher- Wiener Rosenfield ISBT
Race
D
C
E
c
e
14
5
Fisher- Wiener Rosenfield ISBT
Race
D Rh0 Rh1 004001
C rh’ Rh2 004002
E rh’’ Rh3 004003
c hr’ Rh4 004004
e hr’’ Rh5 004005
14
6
Most D>c>E>C>e Least
14
7
 Chr 1
• RHD gene= D polypeptides
• RHCE gene= RHCe, RHcE, RHce, RHCE polypeptides
 Chr 6
• RHAG= Rh-associated glycoprotein that forms
complexes with Rh polypeptides
14
8
FISHER-RACE WIENER
Gene Antigen Gene Agglutinogen Blood factors
Dce Dce R0 Rh0 Rh0 hr’ hr’’
DCe
DcE
DCE
dce
dCe
dcE
dCE
Note:
R or r = D or d
1 or ‘ = Ce
2 or ‘’ = cE
Z or y = CE 14
9
Wiener Fisher-Race Caucasian Black Native Asian
American
R0 Dce 0.04 0.44 0.02 0.03
R1 DCe 0.42 0.17 0.44 0.70
R2 DcE 0.14 0.11 0.34 0.21
Rz DCE 0.00 0.00 0.06 0.01
r ce 0.37 0.26 0.11 0.03
r’ Ce 0.02 0.02 0.02 0.02
r’’ cE 0.01 0.00 0.01 0.00
ry CE 0.00 0.00 0.00 0.00
15
0
Phenotype Equivalent
G D+ C+
f ce (cis)
rhi Ce
Cw
Rh:13; Rh:14; Rh:15; Rh:16
Hr0
Rh:23, Rh:30, Rh:40
Rh:33
Rh:32
e variants
V, VS
15
1
 Weakened form of the D antigen
 D-negative donors and obstetric patients
must be tested for Du
 Du positive donors are classified as D(+)
 Du positive recipients should always
receive D(-) blood
15
2
 Mechanisms
• Genetic weak D
 Complete expression of D antigens but few in number
 Frequent in blacks
• C Trans
 Position effect resulting to steric hindrance
 Dce/dCe ( D and C in trans)
• Partial D (D mosaic)
 One or more of the D epitopes within the entire D
protein is either missing or altered
15
3
-De -DE
CD- cD-
-D-
15
4
D-deletion genes (D● ●)
Rh Deleted
• D--/D--
• No C/c and E/e antigens
• Unusually strong D antigen expression
15
5
 Amorphic type
 Lack all Rh antigens including Rh29
• Rh29 is the highest incidence Rh antigen present
in all RBCs except Rhnull individuals)
 ---/---
 Stomatocytes + Compensated hemolytic
anemia
15
6
 Convert R1R2 to Fisher-Race
15
7
 Px R1R1 ; Donor R2R2. What antibody
can be produced?
15
8
 Reactionof Px RBC with the following
anti-sera?
Anti-D +
Anti-C +
Anti-E -
Anti-c -
Anti-e +
Genotype of Px?
a. rr’ b. r’r’ c. R1/R2 d. R1R1
15
9
 Reactionof Px RBC with the following
anti-sera?
Anti-D +
Anti-C +
Anti-E +
Anti-c +
Anti-e +
Genotype of Px?
a. r’r’ b. r’r’’ c. R1R0 d. R1R2
16
0
 Mother: R1r
 Father: R0r
R1 r
R0
16
1
 Which of the following reacts with anti-f?
a. R1R1
b. R2R2
c. rr
d. All of the above
16
2
 Rh antigens
Reagent Notes
High-protein IgG anti-D Most commonly used
reagents Require the use of an anti-D control
IgM anti-D reagents Immediate spin saline testing

Not for Du testing
Chemically modified IgG Direct saline agglutination testing

anti-D reagents
Monoclonal anti-D Single clones of antibody-producing

reagents cells
Monoclonal blends Combinations of monoclonal IgM

and IgG
16
3
 Rh positive
 Rh negative
 Rh typing
• RBCs + anti-D  Agglutination
• RBCs + anti-D  No agglutination
Du typing (Test for Weak D)
16
4
 Indirect AHG/ Coomb’s test
• RBC + anti-D  37°C incubation
• (in vitro sensitization)
• Wash 3x + AHG  Agglutination
No agglutination
Note:
Rh negative patients may be given Rh (+) blood provided:
a) No previous exposure to Rh (+)
b) Rh0 gam administration
16
5
 Rh viewbox
45-50°C
RBC + anti-D
(37°C)
Read for
agglutination
within 2 minutes
16
6
 Rh antibodies
• IgG (IgG1 and IgG3)
• Immune antibodies
• React optimally at 37°C or after antiglobulin
testing
• Do not bind complement
16
7
 Rh antibodies
• Enhanced reaction by using enzyme-treated
RBCs
 DAT
 IAT
• Extravascular RBC destruction
• Delayed hemolytic transfusion reacton
• Causes HDN
16
8
 Transfusion Reactions
• Highly immunogenic (D Ag most immunogenic)
• 1° exposure = 120 days
• 2° exposure = 2 to 7 days
• Fever, ↑bilirubin, ↓hemoglobin, ↓haptoglobin
• DAT (+)
• Antibody screen
• Antibody elution
16
9
 Rh Hemolytic Disease of the Newborn
• Severe HDN
• No complement activation
• Extravascular hemolysis
• Mother:
• Father:
• Baby:
17
0
• Rh Ig (Rh0 gam)
 Purified anti-D (Artifical Passive Immunity)
 Administered within 72 h after delivery of first child
1. FULL DOSE
• 300 μg anti-D
• Protects up to 30mL D+ whole blood or
15mL D+ RBCs
• >12 weeks gestation
17
1
2. MINI OR MICRO DOSE
• 50 μg anti-D
• Protects up to 5mL D+ whole blood or
2.5mL D+ RBCs
• <12 weeks gestation
17
2
# of RhIg vials = vol of fetomaternal hemorrhage (FMH)
30
# of RhIg vials +1
Vol of FMH= % fetal cells x 50
Problem:
Kleihauer-Betke test reveals 3% fetal cells. How

many vials of Rhogam will be administered?
17
3
3 Rh Blood Group System
17
4
17
5
 Serum antigen secondarily absorbed to
the red cells
 Le gene produces Lea
 Secretors change the Lea to Leb
 Le may also modify the A antigen
review the relationship to ABO
precursors
17
6
 Legene codes for the production of
fucosyltransferase enzyme
• Catalyze addtion of fucose to the 4th C of N-
acetylglucosamine of Type I precursor
substances
17
7
 Legene codes for the production of
fucosyltransferase enzyme
• Catalyze addtion of fucose to the 4th C of N-
acetylglucosamine of Type I precursor
substances
Chromosome 19
17
8
Genes Lewis Red Cell Phenotype
Le Se Lea- Leb+
Le se Lea+ Leb-
lele Lea- Leb-
le se Lea- Leb- Lec+
le Se Lea- Leb- Lec- Led+

17
9
Genotype Substance Red Cell Lewis
(Secretion) Phenotype Abs
ABH, lele,
sese
ABH, lele,
SeSe or Sese
ABH LeLe or
Lele, sese
ABH, LeLe or
Lele, SeSe or
sese
18
0
Genotype Substance Red Cell Lewis
(Secretion) Phenotype Abs
ABH, lele, None ABH Le (a-b-) Anti-Lea
sese Anti-Leb
ABH, lele, ABH ABH Le (a-b-) Anti-Lea
SeSe or Sese Anti-Leb
ABH LeLe or Lea ABH Le (a+b-) Anti-Leb
Lele, sese
ABH, LeLe or ABH, Lea, ABH Le (a-b+) None
Lele, SeSe or Leb
sese
18
1
Lea  Leb
Se
 Produced by tissue cells
 Not well developed at birth
 Decrease in expression of Lewis antigens
has been demonstrated from many
pregnant women
18
2
Newborns born Le a-b-
If Le and Se
• 2 weeks to 6 months Le a+
• then Le a+b+
• then Le a-b+
During pregnancy, antigens become weaker
18
3
Phenotype White Black
Le a+b- 22% 23%
Le a-b+ 72% 55%
Le a-b- 6% 22%
18
4
 Anti-Le a, Anti-Le b, Anti-Lex
 Naturally occurring, IgM
 Most react at room temperature but
may react at 37°C
 Often fix complement
 Some cause in vitro hemolysis
 Le a may cause HTR
18
5
 Anti-Le a
 Found in Lea-b- secretors
 best room temperature or below -
some at ICT and enzymes
 Often fix complement
 Some in vitro hemolysis
 Le a may cause HTR
18
6
 Anti-Le b
 Often found with Anti-Lea
 Most react at room temperature or
below
 Two types - Anti-LebH and Anti-LebL
 Rare cause of HTR
18
7
 Anti-Lex
 Most react at room temperature or
below -
 Reacts with both Lea and Leb as a
single antibody
18
8
 Special Problems in the Blood Bank
• Lewis antigens may be weaker during
pregnancy and women produce antibodies
• Can neutralize Lewis antibodies with Lewis
plasma
• Pregnant woman with room temperature
antibodies, neutralize with Lewis antigen
when testing for HDN antibodies
18
9
Lewis substance can be used to
neutralize the antibodies and
allow the detection of any other
antibodies present
19
0
 Many antigens in this system and some
are alleles to the four common antigens
 M N S s
 Association with GPA and GPB
 Four gene complexes
MS Ms NS Ns
 Other alleles Mg, Mk, Mc, Mr, Mz, Mv, Na, T1m, Sj,
S2, some quantitative differences
19
1
 MN Antigens
• Found on Glycophorin A (MN-Sialoglycoprotein)
• MN antigens differ in their amino acid residue at
positions 1 and 5: M has serine and glycine
whereas N has leucine and glutamic acid
• Well developed at birth
• Destroyed or removed by enzymes
• Paternity testing of infants and young children
19
2
 Ss Antigens
• Found on Glycophorin B (Ss-Sialoglycoprotein)
• Amino acid at position 29 on GPB is critical to
antigen expression: S has methionine whereas s
has threonine
• Less easily degraded by enzymes
19
3
U antigen is absent or reduced on S-s-
 Mi - abnormal forms of Ss glycoprotein
 En(a-) absence of MN glycoprotein
19
4
 Phenotypes
19
5
 Anti-M and Anti-N
• Usually room temperature
• IgM saline reaction
• Dosage (antibodies react better with
homozygous cells)
• Destroyed by enzymes
• Possible HDN and HTR if reaction at AHG
19
6
 Anti-M and Anti-N
• Anti-M: some are pH dependent (best at pH
6.5) and glucose dependent
• Anti-Nf: found in dialysis patients on
equipment sterilized with formaldehyde
 Anti-N like antibody
19
7
 Anti-S
• Usually IgM and reacts at room temperature
although some at AHG
• Rare HTR and HDN
19
8
 Anti-s and anti-U
• Usually IgG and reacts at AHG
• Not destroyed by enzymes
• HTR and HDN
• Anti-U found as warm autoantibody and
does not react well with Rh null cells
• Other antibodies rarely detected but not
uncommon (ex. anti-Mg common antibody)
19
9
 Discoveredin 1927 by Landsteiner
 Antigens P1 P p pk Luke
(p null)
20
0
 Found on fetal red cells as early as 12
weeks, but it weakens with gestational
age
 Deteriorates rapidly on storage
20
1
 P1-like antigen has been found in
plasma, droppings of pigeons and
turtledoves as well as in egg white of
turtledoves
 P1 substance has been identified in
hydatid cyst fluid, extracts of
Lumbricoides terristris (common
earthworm) and Ascaris suum
20
2
 Anti-P1 Anti-P Anti-pk Anti- P + P1 + pk
• Anti-P1 - P2 or P2k individuals
 Usually IgM, reacts at room temperature
and saline
 May attach complement
 rarely a problem with transfusion
 easily inhibited with P1 substance
20
3
 Anti-P1 Anti-P Anti-pk Anti- P + P1 + pk
• Anti-P1 - P2 or P2k individuals
 Strong anti-P1 was observed in
individuals infected with Hydatid cyst
disease (_______________________)
 Associated with fascioliasis, Clonorchis
sinensis and Opisthorchis viverrini
infections
20
4
• Anti-P
 found in sera from Pk individuals - an
IgM hemolytic antibody that is clinically
significant
 also found as an IgG biphasic antibody in
paroxysmal cold hemoglobinuria
called Donath-Landsteiner antibody
20
5
• Anti-pk and Anti P + P1 + pk
 Anti-pk has only been found as part of
other antibodies
 Anti-P + P1 + pk found in p individuals
- formerly called Anti-Tja and very
hemolytic
 Isolated by selective adsorption with P1
cells
 P1 individuals with biliary cirrhosis and
autoimmune hemolytic anemia
20
6
 Two antigens I and i
 I antigen present on almost all healthy
adults
 Rare adults that are I negative
 I antigen varies in strength on adult
cells
20
7
 Newborns do not have much I antigen
 Newborns have i antigen
 At about 18 months: i is replaced by I
 Some transitional antigens
20
8
Age group Phenotype
Neonates I (-) i (+)
Adults (18 mos onwards) I (+) i (-)
Note:
i Adult or I (-) phenotype: Herediatry
erythroblastic multinuclearity with positive
acidified serum test (HEMPAS)
20
9
I substance can be found in saliva and
human milk and on lymphocytes and
platelets
 During disease, the I antigens may
alter
21
0
Anti-I anti-i
• Anti-I
 usually reacts at room temperature, saline or
below
 often attaches complement
 doesn’t cause hemolysis unless it reacts at 37oC
 Can be found in almost all sera in low titers and
titers increase during some diseases (viral
infections - syphilis - atypical pneumonia)
 COLD AUTOAGGLUTIN
21
1
• Benign Anti-I
 Normal healthy individuals
 Not associated with in vivo hemolysis
 Weak naturally occuring saline reactive IgM
Ab
 Interferes with reverse typing
 Usually reacts only at 4°C
21
2
• Pathologic Anti-I
 Potent IgM agglutinins with higher titers and
broader thermal range (up to 30-32°C)
 Attach in vivo and cause autoagglutination and
vascular occlusion (Raynaud’s phenomenon) or
intravascular hemolysis
 Cold agglutinin disease (CAD)/ Cold agglutinin
syndrome (CAS)/ Cold hemagglutinin disease
(CHD)
 Primary atypical pneumonia (________________)
21
3
 Autoanti-I
 Patients with Mycoplasma pneumoniae
often develop strong cold agglutinins with I
specificity as a cross-reactive response to
Mycoplasma antigen
 Listeria monocytogenes organism from
patient with Cold AIHA has been reported
to absorb anti-I and stimulate its
production in rabbits
21
4
• Anti-i
 Mostly IgM
 Rare antibody occurs in patients with:
 Infectious mononucleosis (_______)
 Diseases of the RES:
 Cirrhosis
 Myeloid leukemia
 Reticulosis
21
5
• Other combination antibodies have been
found (IA, IH, IP1, etc.
• ENZYMES ENHANCE ACTIVITY
• ABSORPTION IS USED TO TEST FOR OTHER

MORE IMPORTANT ANTIBODIES
21
6
21
7
Adult Cells Cord Cells
Anti-I
Anti-i
21
8
 Many antigens in this system and has
been given a numerical nomenclature
Refer to table 8-8
 Six most important
Numeric Alpha Name Incidence__________

KEL 1 K Kell 10%
KEL 2 k Cellano 99.8%
KEL 3 Kpa Penny 2%
KEL 4 Kpb Rautenberg 99.9
KEL 6 Jsa Sutter Rare (19% Blacks)
KEL 7 Jsb Matthews 99.9%(99.8% Blacks)
21
9
 Most common gene complexes
22
0
 Immunogenic, K is rated second only to D
in terms of immunogenicity
 Well developed at birth
 Synthesized on precursor, Kx
 Found on WBCs and RBCs
• WBCs: unconverted
 If absent: Chronic granulomatous disease (CGD)
• RBCs: converted to Kell antigens
 If absent: MacLeod phenotype
22
1
 Mc Leod syndrome
• Reduced expression of Kell antigens
• Association with hemolytic anemia and
chronic granulomatous disease
• Acanthocytes
22
2
 Usually IgG and require AHG
 Rare reaction in saline
 Common antibodies
 Implicated in HTR and HDN
 Anti-K is a very common antibody
22
3
 Anti-K
• IgG antibody reactive in the antiglobulin phase
• Immune antibody made in response to Ag
exposure through pregnancy and transfusion
• Severe HTR
• Severe HDN
• Extravascular hemolysis
22
4
 Antibodiesto Kpa, Jsa and other low
frequency Kell antigens
• Rare because so few people are exposed to the
antigen
• Detected through unexpected incompatible
crossmatches or cases of HDN
22
5
 Antibodiesto k, Kpb, Jsb and other high
frequency Kell antigens
• Rare because so few people lack the antigen
22
6
 Discovered in early 1950’s
 Fy antigen locus on chromosome 1
with Rh locus
 Antigens
codominant inheritance
• Fya Fyb Fyx
• Others Fy3 Fy 4 Fy5 Fy6 Fs - (page
185)
22
7

22
8
Mr. Duffy
• Multiple transfused hemophiliac
• Fya (1950)
 Serum of a woman with three
pregnancies
• Fyb
22
9
Fy (a+b+): common among Whites
Fy (a+b-): common among Chinese
Fy (a- b-) common among Blacks
• Resistant to Plasmodium vivax and
Plasmodium knowlesi
23
0
 Fya-b-
appear to provide some
protection from P. vivax infection
• Fy6: important for invasion of P. vivax
23
1
Fya and Fyb
• Identified on fetal red cells as early as 6
weeks gestational age and are well
developed at birth
• Destroyed by common proteolytic enzymes
23
2
 Anti-Fya and Anti-Fyb
• Usually AHG reaction - IgG
• Enhanced by low ionic strength medium
• Rare examples of antibodies to other antigens
(Anti-Fy 3, Anti-Fy4, Anti-Fy5) and those reactions
are not destroyed by enzymes
• Cause HTR and HDN
23
3
 Discovered in the 1950s
 Two antigens Jka and Jkb
23
4
Mrs. Kidd
• Whose infant has HDN
23
5
 Jka and Jkb
• Detected on fetal red cells as early as 11 weeks
for Jka and 7 weeks for Jkb
• Not altered by enzymes
23
6
Anti-Jka and Anti-Jkb
• Notorious reputation in the blood
bank since it is not easily detected
• Usually IgG and require AHG
• Bind complement
• Enhanced by enzymes
23
7
• Implicated in HDN and HTR
• Seldom potent and deteriorate rapidly
• Classic delayed HTR
• Drug-induced hemolytic anemia
 Patients receiving methyldopa
23
8
Anti-Jk3
• found in some Jka-b- individuals
• reacts with Jka and Jkb
23
9
 Two antigens Lua (8%) Lub (99%)
 Important blood group that demonstrates
multiple methods for inheritance of the null
cell type
 Lu a-b- inheritance
• InLu dominate inhibitor gene
• lulu recessive lack of Lu gene
• sex linked inhibitor gene
24
0
Luteran
• Donor of a patient with Lupus
Erythematosus Diffusus who
contained a certain antibody
24
1
 Lua and Lub
 Poorly developed at birth and do not
reach adult levels until age 15
24
2
Antibodies
• Anti-Lua - not common - reacts in saline but
can be IgG and require AHG - gives a (mf)
agglutination - unclear about HTR & HDN
• Anti-Lub - rare - mostly IgG and requires

AHG - probable HTR and HDN
24
3
• Anti-Luab (Anti-Lu3 ) - reacts with all but
Lu a-b- of the recessive type
• Other antibodies react with rare Lu

phenotypes found on Lua-b-
24
4
Blood Antibody Optimum Temp Reaction Enzyme
Group Class Phase Treatment
Kell IgG 37°C AHG No effect
Duffy IgG 37°C AHG Destroyed
Kidd IgG 37°C AHG Enhanced
Lutheran Lua IgM Lua 4°C Lua RT Enhanced
Lub IgG Lub 37°C Lub AHG
Lewis IgM Most often 4°C, RT, 37°C, AHG Enhanced
sometimes 37°C
I IgM 4°C IS and occ. Enhanced
37°C
P IgM (anti-P1) 4°C IS, 37°C, AHG Enhanced
MNS
MN IgM 4°C or 37°C IS, 37°C, AHG Destroyed
Ss IgG 37°C AHG Variable 24
5
4
Other Blood Group Systems
24
6
24
7
 Diego - Dia Dib Wra Wrb 3 others
• Dia found in Chippawah Native Americans
and Japanese and Chinese
• Dia useful tool in anthropologic studies of
Mongolian ancestry
• Uncommon antibodies - AHG reaction and
important in HTR and HDN
• Wra is a low incidence antigen and Wrb is a
high incidence antigen
• Anti-Wra is a fairly common antibody - IgM or
IgG
24
8
 Diego - Dia Dib Wra Wrb 3 others
• Low incidence antigens: Wda, Rba and WARR
• Defect in Anion Exchange Protein (AE-1) also
known as erythrocyte band 3
 Hereditary spherocytosis
 Congenital acanthocytosis
 Southeast asian ovalocytosis
24
9
Chido/Rogers
• Nine antigens - all normal individuals are
either Rg + or Ch +
• HTLA - use plasma inhibition
• Determinants on C4 molecule and linked
to HLA
25
0
Xg
• No known antithetical partner
• Sex-linked inheritance
 Xga positive Male - 66% Female - 89%
• Uncommon antibody - AHG reaction and
destroyed by enzymes - HTR and HDN?
25
1
 Gerbich
• system with at least 3 high incidence antigens
(Ge2, Ge3, Ge4) and 4 low incidence antigens
(Wb, Lsa, Ana, Dha)
• Inherited on Chr 2 and expressed on
glycophorins C and /or D
• Antibodies usually IgG which require AHG
and clinically significant
25
2
Scianna
• Sc:1 - 100% Sc:2 - 0.3% Sc:3 - 100%
• Antibodies are rare
25
3
Colton
• Colton antigens:
 Coa-99.7% Cob 10.7% Co3 (Coa)100%
• Co antigens have been located on the
transport protein known an channel-
forming integral protein (CHIP)
• Null phenotype has been found and
associated with genetic abnormality and
anemia
• Antibodies: IgG and clinically significant
25
4
Cromer
• Consists of 7 high incidence antigens and
three low incidence antigens
• Antigens are carried by decay accelerating
factor (DAF) which is involved in the regulation of
complement activation by accelerating decay of
C3 and C5 convertase
• Anti-CROM antibodies (black pop)
probably clinically significant
25
5
Cartwright
• Antigens Yta - 99.8% Ytb - 0.2%
• Yt antigens have been located on
erythrocyte acetylcholinesterase, an
enzyme involved in neurotransmission
• Usually IgG and detected by AHG
• ?HDN and HTR?
25
6
Dombrock
• Antigens Doa - 57% Dob - 83%
• Additional antigens added Holly, Gregory,
and Joseph
• Uncommon antibodies HTR and ?HDN?
25
7
 Indian
• Ina Inb
• Ina Iranian and Arabs
• IN antigens are carried on the hematopoietic
isoform of the CD44 marker, which is known
for its immune adhesion properties
• IN antibodies: IgG, red cell stim., IAT
• Enzyme destroyed - Ina HTR
25
8
Knops
• Five antigens: Kna, Knb, McCa (McCoy), Sla
and Yka (York)
• Alleles on Chr 1 with Ag residing in
complement receptor 1 (CR1)
• depressed in some diseases
• HTLA
• AntibodiesL IgG, red cell stim., weak and
var. rxn at IAT, ?HDN and ?HTR
25
9
HLA antigens detectable on RBCs
• Bga with HLA B-7
• Bgb with HLA B-17
• Bgc with HLA A-28
• Antibodies: IgG, weak and var in IAT,
?HDN and ?HTR
26
0
 Exhibit
reactivity at high dilutions of
serum but the strength of agglutination
is weak at any dilution
Anti-Ch Anti-Rg Anti-Kn Anti-JMH
Anti-Yka Anti-Csa Anti-McC
26
1
5 Minor Blood Group Systems
26
2
26
3
1. Appears to be in good health
2. Age
• AABB: 17 years old
• PH: 18-65 years old
• <18 years old (16, 17 y/o)= Parent’s consent
• >65 years old = Physician’s consent
26
4
3. Body weight
• At least 110 lbs/ 50 kg
 450mL blood (63mL anticoagulant)
 30mL (tubing) for serologic tests
• Maximum of 10.5mL blood/ kg body weight
26
5
3. Body weight (cont)
• Volume of blood to be drawn
 (Donor’s wt/ Ideal wt) x 450
• Volume of anticoagulant needed
 (Vol of blood to be draw/100) x 14
• Volume of anticoagulant to be removed
from the blood bag
 63- Vol of Ac needed
26
6
 Note:
• Blood bag
 63mL 450 mL blood
 31.5 mL200 mL blood
• 1 part Ac : 7 parts blood
• Coagulation studies
 Light blue top
 1 : 9 (Ac: blood)
• Westergren ESR
 Black top
 1:4 (Ac: blood)
26
7
4. Temperature
• Oral Temperature
 ≤37.5°C or 99.5°F
5. Pulse
• 50 to 100 bpm
• Lower pulse rate for athletes
26
8
6. Blood pressure
• A. AABB
Systole not to exceed 180mmHg
Diastole not to exceed 100 mmHg
• B. Philippine setting
Systole: 90-160 mmHg
Diastole: 60-100mmHg
26
9
6. Blood pressure
• CuSO4 method
 1.053 SG
 1 cm distance from
blood drop to solution
 15 seconds
 30mL container
 25 tests
 Changed daily
27
0
7. Hemoglobin and Hematocrit
Autologous Allogeneic
Hemoglobin
Hematocrit
27
1
7. Hemoglobin and Hematocrit
Autologous Allogeneic
Hemoglobin ≥11 g/dL ≥12.5 g/dL
Hematocrit ≥33% ≥38%
27
2
1. Homologous/ Allogeneic
 Voluntary donors of blood for the national
supply
2. Autologous
 One who donates blood for his or her own
use
a) Preoperative collection
b) Acute normovolemic hemodilution
c) Intraoperative collection
d) Postoperative collection
27
3
27
4
3. Directed
 Same requirements as allogeneic except
that unit is directed towards a specific
patient
4. Pheresis
 Donor in which blood is withdrawn from a
donor or patient and separated into its
components with one or more of the
components retained and the remaining
returned to the individual
27
5
 Hemapheresis
 Gk. meaning “to separate or remove”
 45-120 minutes
1. Plateletpheresis (Thrombocytapheresis)
2. RBC pheresis (Erythrocytapheresis)
3. WBC pheresis (Leukapheresis)
4. Plasmapheresis
5. Stem cell pheresis
27
6
27
7
1. Intermittent flow centrifugation
 Requires only one venipuncture
 Blood is withdrawn and reinfused
through the same needle
 Once separated, the remaining
components are reinfused and one
cycle is completed
27
8
2. Continuous flow centrifugation
 Two venipuncture sites are necessary
 Withdraw, process and return the blood
to the individual simultaneously
27
9
 Anticoagulant: ACD (commonly),
heparin
 Prime the system: NSS
28
0
 Procedure: 45 to 90 minutes
 Every 2 days (not to exceed twice a week)
 No aspirin or NSAIDs for 3 days
 Storage:Open system- 24 h
(20-24⁰C) Closed system-5 days
 QC: 3.0x1011
 Equivalent to 6 to 10 Random Platelet

28
1
 Sedimenting agent: Hydroxyethyl starch
• Heavier density
• WBC settle
 Corticosteroids, G-CSF, GM-CSF
 Storage: 24 hours (20-24°C)
 1x1010 granulocytes
28
2
 Double RBC pheresis
 Every 16 weeks
Sex Hct Weight Height

Male 40% 130 lbs 5’ 1’’
Female 40% 150 lbs 5’ 5’’
28
3
 Once every 4 weeks
 Replacement fluid (plasmapheresis):

5% NSA (commonly), NSS
28
4
1. Donor Registration
2. Interview and Physical Examination

• Multiple needle marks
• Unexplained weight loss
• Oral thrush and Kaposi sarcoma
28
5
3. Actual donor selection and
blood collection
• 2 blood bank personnel
 1 Med Tech and 1 head of blood bank
• Donor bleeding
 7-10 minutes (AABB)
 ≤15 minutes (PH)
 >15 minutes= not for cryoprecipitate
28
6
blood collection
• Aseptic technique
 PVP-iodine or polymeriodine complex
 Chlorhexidine gluconate and isopropyl alcohol
• 45° angle and reduce to 10-20° once in
punctured in the skin
28
7
blood collection
• 45° puncture angle and reduce to 10-20°
once in the skin
• Sphygmomanometer as tourniquet:
 40-60 mmHg
28
8
Permanent
• High risk history for AIDS
 Men who have had sex with another man any time
since 1977
 Hemophiliacs
 IV drug abusers (past or present)
 Persons who have engaged in sex for money or
drugs anytime since 1977
 Confirmed laboratory test for AIDS
28
9
Permanent
• Symptoms of viral hepatitis after age 11
• Confirmed positive test for HBsAg, anti-HBc
• Confirmed positive test for HC Ab
• Confirmed positive test for Human T-cell
lymphotropic virus (HTLV ½)
• Malignant solid tumors except:
 Basal cell carcinoma of the skin
 Carcinoma in situ of the cervix
29
0
Permanent
• Hematologic malignancies
• Chemotherapeutic agents administered for
malignancy
• Chronic cardiopulmonary, liver or renal
disease
• Serious abnormal bleeding tendencies
• Intake of drug etretinate (Tegison) for
treatment of psoriasis
29
1
Permanent
• History of babesiosis, Chagas’ disease
• Recipient of pituitary derived growth
hormone
• Recipient of cornea/ dura mater transplant
29
2
Temporary
• Active disease under treatment such as
cold, flu, tuberculosis, syphilis, infections,
curable disease of the heart, lung, kidney,
liver and gastrointestinal tract and
treatment of antibiotics
29
3
3 year deferral
• After departure of an immigrant or refugee coming
from an area endemic for malaria
• After being asymptomatic for those who have been
diagnosed with malaria
29
4
1 year deferral
• After hepatitis B immune globulin administration
• After therapeutic rabies vaccination
• Rape victims
• Health care workers with percutaneous exposure to
blood or body fluids
• Close contact with viral hepatitis
• Tattoo
• Sexual contact with a prostitute or other persons in a
high-risk group for AIDS
29
5
1 year deferral
• Incarceration in a jail for more than 72 consecutive
hours
• Major blood transfusion
• Major surgical operation
• Travel to areas endemic for malaria (with or without
prophylactic therapy)
• History of syphilis or gonorrhea (Under treatment or
positive screening test)
29
6
AABB PH
Two months Three months (12 weeks) after
after recent recent blood donation [450 mL]
blood donation
Six to eight weeks after recent
blood donation [200mL]
Six weeks Nine months
after delivery after childbirth
of a baby
29
7
Four weeks (1 month) Two weeks
After vaccination with:
German measles (rubella) Oral polio, measles
(rubeola), mumps,
yellow fever,
immune reaction to
smallpox
After cessation of drug:
Isoretinoin (Accutane) for acne
Finasteride (Proscar) for BPH
29
8
3 months
• After typhoid fever infection
1 month (4 weeks)
• After household contact with typhoid
2 months (8 weeks)
• After MMR vaccination
2 to 3 weeks
• After febrile episode
3 days (if for use of platelet components)
• After intake of aspirin
2 days (if for whole blood donation)
• After hemapheresis
 12 to 24 hours
• After recent alcohol intake 29
9
Common Donor Attributed Prevention/
Reactions to Management
Lightheadedness Anxiety Reassuring
Weakness Hypoglyc. conversation, elevate
Tingling sensation donor’s feet, apply
Palpitations cold, wet towels to
neck and forehead,
have donor breathe
into paper pag,
provide juice before
donation
Discontinue donation300
Fainting Anxiety Discontinue donation
Hypoglyc. Administer glucose
solution
Provide juice before
donation
Position donor in
place protected from
possible fall
30
1
Convulsion Anxiety or Discontinue donation
underlying Elevate feet
Restrain gently to
disease
prevent injury
Maintain airway
Reassure after
recovering
consciousness
Inform about possible
involuntary loss of
control of urine and stool30
2
Cardiopulmonary Underlying Ventilation
emergency heart CPR (if necessary)
disease Transfer donor to
emergency medical
facility
30
3
Common Donor Attributed to Prevention/
Reactions Management
Hematoma Very fragile Discontinue if
veins; large
Unskilled Apply pressure to
phleb or site for at least 5
uncooperative minutes
donor Apply cold packs
Reassure donor
30
4
Jet-like pulsating Inadvertent Discontinue ASAP
bleeding with puncture of Apply firm
bright red blood artery when pressure over
deep vein puncture site for at
punctures are least 10 minutes
attempted Apply dressing on
site
Follow-up donor
for additional care30
5
Shooting pain Inadvertent Reassure
followed by puncture of Apply support to
numbness and median nerve the arm
tingling in the or cutaneous
forearm branches
(rare)
30
6
6 Blood Donors
30
7
30
8
Component Mechanism of action
Citrate
Dextrose
Citric acid
Phosphate buffer
Adenine
30
9
Component Mechanism of action
Citrate Binds calcium
Dextrose Provide red cell energy
Citric acid Lowers pH preventing
caramelization
Phosphate buffer Raise ATP
Adenine Improve RBC survival
31
0
Acid-citrate-
dextrose (ACD)
Citrate-phosphate-
double dextrose
(CP2D)
Citrate-phosphate-
dextrose (CPD)
Citrate-phosphate-
dextrose-adenine
(CPDA-1)
CPDA-2 31
1
Acid-citrate- 21 days
dextrose (ACD)
Citrate-phosphate- 21 days
dextrose (CPD)
double dextrose
(CP2D)
dextrose-adenine
(CPDA-1)
CPDA-2 42 days 31
2
1. Saline- solute is suspended
2. Adenine- improved RBC
survival
3. Glucose- provide RBC energy
4. Mannitol- RBC membrane
stabilizing agent
31
3
Additive solutions
Adsol (AS-1)
Nutricel (AS-3)
Optisol (AS-5)
31
4
Additive solutions
Adsol (AS-1) 42 days
Nutricel (AS-3) 42 days
Optisol (AS-5) 42 days
31
5
 Regenerate ATP and 2,3-DPG
 Red cells stored in the liquid
state for fewer than 3 days after
their outdate are rejuvenated
for 1 to 4 hours at 37°C
31
6
 Rejuvesol = FDA approved
 PIGPA
• Phosphate, inosine, glucose,
pyruvate adenine
 PIPA
• Phosphate, inosine, pyruvate
adenine
31
7
pH
ATP
2,3-DPG
Plasma hemoglobin
Plasma potassium
Plasma sodium
31
8
pH ↓
ATP ↓
2,3-DPG ↓
Plasma hemoglobin ↑
Plasma potassium ↑
Plasma sodium ↓
31
9
Whole blood
5000xg for 5 Hard/ Heavy Light/ Soft 2000xg for 3

minutes (1-6°C) Spin Spin minutes(1-6°C)
Platelet-rich
Plasma pRBCs
plasma
≤-18°C
Fresh Frozen
pRBCs
Plasma
32
0
Fresh Cryoprecipitate
Heavy
Plasma Frozen
Heavy Spin
WB Plasma
spin Cell-free plasma
pRBCs
5000xg for 7
minutes (1-6°C)
32
1
Whole blood
5000xg for 5 Hard/ Heavy Light/ Soft 2000xg for 3

minutes (1-6°C) Spin Spin minutes(1-6°C)
Platelet-rich
Plasma pRBCs
plasma
≤-18°C
Fresh Frozen
pRBCs
Plasma
32
2
5000xg for 5
minutes (1-6°C)
Platelet
Plt-rich Heavy concentrate
Light Plasma Spin Platelet-poor
WB FFP Cryoppt
spin plasma
pRBCs
32
3
 Centrifugation
 Heavy Spin
• 5000xg for 5 mins (pRBC, plt conc)
• 5000xg for 7 mins (cryoppt, cell-free plasma)
 Light Spin
• 2000xg for 3 mins (plt rich plasma)
 Note:
• Components processed within 6 to 8 hours
• Components are prepared using a refrigerated
centrifuge (1-6°C) except:
 Platelets: RT (20-24°C)
32
4
Component
Indication
Storage
Transport
Shelf-life
Dosage
Quality Control
32
5
Whole Blood
Indication ↑Vol and RBC mass
Storage 1-6°C
Transport 1-10°C [30 mins]
Shelf-life ACD
CPD
CP2D
CPDA-1
CPDA-2/S
Heparin=2 days (48h)
32
6
Packed RBCs
Indication ↑RBC mass
Storage 1-6°C
Transport 1-10°C
Shelf-life Open system: 24 h
Closed system:
ACD, CPD, CP2D
CPDA-1
Dosage ↑Hb 1g/dL; ↑Hct 3%
QC ≤80% Hct
32
7
Leukocyte-Reduced RBCs
Indication ↑RBC mass in Px with severe and or
recurrent febrile NHTR due to
leukocyte Abs; Prevent CMV
Methods Filtration, centrifugation, saline washing
Storage 1-6°C
Transport 1-10°C
Closed system: Same as WB
QC ≤5x106; >85% RBC recovery 32

8
Washed RBCs
Indication ↑RBC mass in Px with allergic,
anaphylactic, febrile and urticarial
reactions
Storage 1-6°C
QC 70-80% Hct
32
9
Frozen RBCs
Indication Storage of rare blood and autologous
units
Storage -65°C or -120°C
Shelf-life 10 years
Notes: Cryoprotective agent- prevent RBC
rupture during freezing e.g. glycerol
33
0
Frozen RBCs
Methods of freezing red cells
Method Component Freezing temp Storage temp
High Glycerol Slow freezing -80°C -65°C
40%w/v (Mechanical
glycerol freezer)
Low Glycerol Fast freezing -196°C -120°C (Liquid

20%w/v nitrogen)
glycerol
Agglomeration Glycerol, -80°C -65°C

glucose, (Mechanical
fructose, EDTA freezer)
33
1
 Deglycerolization
• Removal of glycerol
• Washing with hypertonic solution followed by
isotonic solution
• Use RBCs within 24 h after deglycerolization
Method
High glycerol 12% NaCl 1.6% NaCl  0.9% NaCl
Low glycerol 45% NaCl in 15% mannitol  0.9% NaCl
Agglomeration 50% glucose + 5% fructose  0.9% NaCl
33
2
Platelets
Random Donor Single Donor
Indication Thrombocytopenia, Platelet
DIC, bleeding refractoriness
Storage 20-24C w/ constant agitation
Shelf-life 5 days
Dosage ↑Plt ct 5k-10k/uL ↑Plt ct 30k-60k/uL
QC ≥5.5x1010 Plts ≥3.0x1011 Plts
≥pH 6.2 ≥pH 6.2
33
3
Fresh Frozen Plasma
Indication Correct multiple coagulation
deficiency (e.g. liver disease)
Reverse effects of Warfarin (Coumadin)
Storage ≤-18°C or ≤-65°C
Shelf-life 1 year or 7 years
Notes: Thawed at 37°C
Thawed plasma stored at 1-6°C used
within 24 h
33
4
Cryoprecipitate
Indication Fibrinogen deficiency, hemophilia A, von
Willebrand’s disease and FXIII def.
Storage ≤-18°C
Shelf-life 1 year
Content Fibrinogen: 150 mg, AHF: 80 U, vWF, FXIII
Notes: White mass of precipitate in 15mL plasma
Administer w/in 6h (after thawing)
or 4h (after pooling)
Not harvested if >15 min collection
33
5
Granulocytes, Pheresis
Indication Neutropenia, granulocyte dysfunction e.g.
CGD, myeloid hypoplasia unresponsive to
antibiotics
Storage 20°C-24°C
Shelf-life 24 h
QC 1x1010 granulocytes
Notes: Leukapheresis
Hydroxyethyl starch sedimenting
Adm. Corticosteroids before donation circ.
granulocytes
33
6
Factor VIII concentrate
Indication Prevent or control bleeding in
hemophilia A patients
Storage 1-6°C (lyophilized)
Shelf-life varies
33
7
Factor IX concentrate
Indication Prevent or control bleeding in
hemophilia B patients or with specific
factor deficiencies (also contains the
prothrombin complex: II, VII, IX, X)
Storage 1-6°C (lyophilized)
Shelf-life varies
33
8
Normal Serum Albumin
Indication Replace loss of colloids in
hypovolemic shock, severe burns or
for pressure support during
hypotensive episodes
Contents 96% albumin, 4% globulin
Storage 2-10°C
Shelf-life 5 years
33
9
Plasma Protein Fraction
Indication Replace loss of colloids in
hypovolemic shock, severe burns or
for pressure support during
hypotensive episodes
Contents 80-85% albumin, 15-20% globulin
Storage 2-10°C
Shelf-life 5 years
34
0
UV Irradiation
Heating in Liquid State
Heating in Lyophilized Form
34
1
 Usually RBCs and platelets
 Indication:
• Prevent Transfusion-associated Graft vs. Host
Disease (TA-GVHD)
• Inactivate T-cells
 At risk for GVHD:
• Recipient of BM transplant
• Px with congenital immunodeficiency
• Px with hematologic/ oncologic disorder
• Recipient of blood from first degree relative
34
2
 Radiation source
• Cesium (137Cs)
• Cobalt (60Co)
 Quality control
• 25 Gy35 Gy
 Shelf life
• 28 days from irradiation or original expiration
date (whichever comes first)
34
3
 Must be completed within 4 hours
 Assisted by at least 2 nurses
 NSS: only fluid allowed to start an IV line
prior to transfusion
• Not 5% dextrose or ringer’s lactate
 Filter
• Clot screen filter
• Leukocyte depletion filter
 Blood Warmer: Kept at 37C
 Speed of infusion: 200mL blood/hr
• 4mL/ min
• 60 drops/min
34
4
34
5
7 Blood Components
34
6
34
7
Adverse effect that occurs as a result
of administration of blood or blood
components
• Immediate
• Delayed
34
8
Immediate Delayed
Immunologic Immunologic
• Hemolytic • Hemolytic
• Febrile • TA-GVHD
• Allergic • Post-transfusion purpura
• TRALI
Non-immunologic Non-immunologic
• Bacterial contamination • TA-hemosiderosis
• Circulatory overload • Disease transmission
• PCITR
34
9
 Immediate immunologic
Febrile Nonhemolytic
Definition Inc in temp of 1°C or more
associated with transfusion that
cannot be explained by any other
condtion
Cause Anti-leukocyte antibodies
Remedy Leukopoor RBCs

35
0
Allergic
Definition Reaction between recipient antibody
and transfused donor plasma
proteins
Cause Donor plasma with foreign protein
Remedy Washed RBCs
35
1
Anaphylactic
Definition Afebrile reaction that occurs only
after infusion of only few mL of blood
Cause IgA deficient patient with anti-IgA
Remedy Washed RBCs
35
2
Transfusion-related acute lung injury (TRALI)/
Non-cardiogenic pulmonary edema (NCPE)
Definition Attributed to the administration of
donor plasma containing high
concentrations of leukoagglutinins
Cause Anti-leukocyte antibodies
Remedy Leukopoor RBC
35
3
 Immediate non-immunologic
Bacterial contamination
Definition Contamination of blood products from donors
with transient bacteremia during phlebotomy,
preparation and processing, thawing
Cause Endotoxins of psychrophilic organisms:
Yersinia enterocolitica, E. coli, Pseudomonas
spp.
Prevention Aseptic techniques, visual inspection
(brownish/purplish discoloration, hemolysis,
clots, murky plasma), infusion within 4 h
35
4
Circulatory overload
Definition Associated with rapid infusion of large
volumes of blood products
At risk Children, elderly patients, cardiac disease

patients
Therapy Therapeutic phlebotomy, O2 therapy,
intravenous diuretics
35
5
Physical or Chemical Induced Transfusion
Reaction (PCITR)
Cause •Mechanical damage (infusion through small
bore)
•Osmotic or chemical damage (addition of
hypo- or hypertonic solutions)
•Thermal trauma (freezing blood without
cryoprotectant or warming above 50°C)
•Citrate toxicity
35
6
 Delayed immunologic
Transfusion-associated Graft vs. Host Disease
(TA-GVHD)
Definition Certain susceptible recipients with
compromised immune systems are transfused
with blood or blood components containing
immunocompetent lymphocytes which engraft
in recipient’s tissue and multiply
Cause Proliferation of T cells that reacts against
foreign tissue of the host recipient
Prevention Irradiated Blood Components
35
7
 Delayed immunologic
Post-transfusion Purpura
Definition Previously immunized patients to platelet
antigens through pregnancy or transfusion
produce antibodies once stimulated,
destroying patient’s own platelets
Cause Platelet antibodies
Therapy Exchange transfusion, corticosteroids,

intravenous immunoglobulin
35
8
 Delayed non-immunologic
TA-hemosiderosis
Definition Iron overload accumulating in the
mitochondria of cells in organs like the liver,
heart and endocrine gland
At risk Patients that are chronically transfused
•Thalassemia major
•Sickle cell anemia
•Hemoglobinopathies
Therapy •Iron chelating agent (Desferrioxamine)
•Neocytes (young RBC with long lifespan)
35
9
 Delayed non-immunologic
Disease transmission
Definition •Hepatitis B, C, D
•CMV
•EBV
HTLV 1 and 2
•HIV
•T. pallidum
•Plasmodium spp.
•B. microti
•T. cruzi
•T. gondii 36
0
 Transfusion must be stopped for any
reaction.
 IV line must be kept open with
crystalloids in case immediate treatment
is necessary to overcome hypotension
 Attending physician and the blood bank
must be notified ASAP
36
1
 Work-up
1. Clerical check (compatibility tag, label,
Px ID)
2. Examination of pretransfusion clotted
specimen, EDTA Ac post-transfusion
blood specimen (perform DAT) and
blood bag
3. Perform GS on blood bag and culture if
necessary
36
2
 Work-up
4. Repeat ABO/Rh, antibody screen and
crossmatch. If antibody is suspected, do
RBC panel
5. Examination of post-transfusion urine
6. Determination on post-transfusion
specimen PT, aPTT, Plt ct, fibrinogen,
FSP
7. Measurement of Hct/Hb frequently
36
3
8 Transfusion Reactions
36
4
36
5
1. Blood typing
2. Compatibility testing
3. Antibody detection and
identification
36
6
Pretransfusion compatibility testing
• Series of testing procedures and processes

with the ultimate objective of ensuring the
best possible results of a blood transfusion
36
7
 Pretransfusion compatibility testing
1. Identification of the patient and donor and

collection of appropriate samples for testing
2. Testing of the donor blood sample
3. Testing of the patient sample and review of
past blood bank records
4. Selection of appropriate donor units
5. Crossmatching
6. Reidentification of the patient before infusion
of blood
36
8
 Major cause of transfusion-associated
fatalities is clerical error
 Recipient’s wristband identification must
always be compared with the requisition
form
 If the patient does not have a wristband
and is coherent, ask the patient to state
his or her full name and to spell it out
36
9
 Serum or plasma may be used for
pretransfusion testing
 Serum is preferred because plasma may
cause small fibrin clots to form
 Plasma may also inactivate complement
so that some antibodies may not be
detected
 Stored for minimum of 7 days following
transfusion at 1-6°C
37
0
ABO/Rh Testing
• Tube method using monoclonal reagents

• Patient sample 3 days old if, within past 3
months, patient has been
 Pregnant
 Transfused
37
1
Historical record check
• Every patient, every time
37
2
Historical record check
• Every patient, every time
37
3
Screening for unexpected
alloantibodies
• Tube method
• RBC, PEG and LISS enhancement media
• Gel method
• Detect clinically significant alloantibodies
reactive at 37°C
• DAT and autocontrol not required
37
4
 Recipient’s serum or plasma must be
tested for clinically significant
unexpected antibodies
 Incubation at 37°C and Coomb’s test
Usually Sometimes Very Unusual

ABO, Rh, Cartwright, Bg (HLA),
Kell, Duffy, Lutheran, Gerbich, Ch/Rg, Leb,
S, s, U, P Dombrock, M, N, JMH, Xga
Lea, Vel, LW, Ii, H,
Ata, Inb, Mia, Csa
37
5
Alloantibody identification
• Tube method with LISS enhancement media

• Gel method on the increase
• Additional methods:
 Prewarm serum
 Increase serum-to-cell ratio
 Enzyme treated reagent RBCs
 PEG
37
6
Once antibody has been detected,
additional testing is necessary
Patient’s history
Reagents
• Antibody identification panel: collection of 11
to 20 group O cells with various antigen
expression
 Profile
sheet specifying the antigens on
each cell and providing a place to record
reactions 37
7
Exclusion
• Rule out antibodies that could not be responsible for
the reactivity seen
• Do exclude only if the antigen is homozygously
expressed on cell to avoid weak antibody showing
dosage
Evaluation
• Remaining antigens should be examined to see if the
pattern of reactivity matches a pattern of antigen-
positive cells
37
8
 Logical approach
• Phase/s and strength of positive reaction
• Positive cells at same phase or at different
phases
• Matched serum reactivity to remaining
specificities
• Rule-out of commonly encountered RBC Ab
• Autologous control (positive/negative)
• Evidence to prove suspected antibody
• Patient lacking the Ag corresponding to Ab
37
9
Cell D C Lea Leb IS 37°C AHG
1 + + + + 0 1+ 1+
2 + + + 0 0 1+ 1+
3 + 0 0 + 0 0 0
4 + 0 0 0 0 0 0
38
0
Cell Fya Fyb Jka Jkb IS 37°C AHG Enzyme
1 + + + + 0 1+ 1+ 4+
2 + + + 0 0 1+ 1+ 4+
3 + 0 0 + 0 0 0 0
4 + 0 0 0 0 0 0 0
38
1
Cell C D E c e K k M N 37°C AHG Check cells
1 + + + + 0 0 + 0 + 2+ 3+ Not perf.
2 0 + 0 + + 0 + + 0 0 0 2+
3 0 + + + 0 0 + + 0 2+ 4+ Not perf.
4 + + 0 0 + + + + + 0 0 2+
5 0 0 0 + + 0 + + 0 0 0 2+
38
2
Coomb’s control cells
(check cells)
• RBCs coated with human IgG antibody
which are added to all AHG-negative
tube tests to ensure that there was an
adequate washing step performed and
that the AHG reagent is present and
functional in the test system
38
3
Cell C D E c e K k M N 37°C AHG Check cells
1 + + + + 0 0 + 0 + 2+ 3+ Not perf.
2 0 + 0 + + 0 + + 0 0 0 2+
3 0 + + + 0 0 + + 0 2+ 4+ Not perf.
4 + + 0 0 + + + + + 0 0 2+
5 0 0 0 + + 0 + + 0 0 0 2+
38
4
Here
38
5
Management of previously identified
alloantibodies
• Honor all clinically significant alloantibodies,

even if currently undetectable
• Unnecessary to reidentify previously
identified alloantibodies
38
6
Crossmatch
• Serologic
 Use immediate spin only if no currently or
historically identifiable
 Use AHG crossmatch if an alloantibody has been
currently or historically identified
• Electronic
 Use only when immediate spin crossmatch would
have been used
38
7
Testing of the patient’s serum with
the donor RBCs including an
antiglobulin phase or simply an
immediate spin phase to confirm
ABO compatibility
Crossmatching is just a part of
compatibility testing!
38
8
 Originally preceded antibody screening
as part of pretransfusion
 Final check of ABO compatibility
between donor and patient
 Detect the presence of an antibody in the
patient’s serum that will react with
antigens on the donor’s RBCs but that was
not detected in antibody screening
because the corrresponding antigen was
lacking from the screening cells
38
9
 Compares recent ABO serologic results
and interpretations on file for both donor
and recipient
 Can replace immediate spin crossmatch
when two blood types are on file for the
patient and antibody screen is negative
39
0
 Major Crossmatch
• Tests recipient serum vs. donor red blood cells
 Minor Crossmatch
• Test the compatibility of the donor's
serum/plasma with the red blood cells of the
recipient
39
1
 Minor crossmatch
• rarely performed, for two main reasons:
 transfused blood is screened for unexpected (non-
ABO) antibodies,
 since the volume of transfused serum is generally
small in comparison to the patient's blood volume,
minor incompatibilities are usually not of great
consequence
39
2
Reaction phase Ig Class Application (aside Type of
Commonly from crossmatching) Antibodies
Detected
Immediate Spin IgM ABO reverse typing Expected ABO
Antibody alloantibodies
screening/identification Unexpected cold
Autocontrol reacting
alloantibodies or
autoantibodies
37C incubation IgG Antibody screening/
identification
Autocontrol
Antiglobulin test IgG Antibody screening/ Unexpected
identification warm-reacting
Autocontrol alloantibodies or
DAT autoantibodies
39
3
39
4
DAT IAT
Principle Detects In-Vivo Detects In-Vitro
RBC Sensitization RBC Sensitization
Application HDN Antibody
HTR detection
AIHA Antibody
identification
Antibody titration
RBC phenotype
39
5
Ratio of serum to cells 40:1 (2 drops serum + 1 drop 5% RCS)
133:1 (4 drops serum + 1 drop 3% RCS)
Reaction medum Albumin: allow antibody-coated cells to come

into closer contact with each other so that
aggregation occurs
LISS: enhance antibody uptake and allow
incubation times to be decreased
PEG: increase antibody uptake by removal of
water to concentrate the antibodies
39
6
Temperature 37 C (optimum temp for IgG and
complement activation)
Incubation time 30 and 120 minutes; LISS= 10 to 15 mins
Washing Min of 3 times saline washing;

FALSE NEG if incomplete due to
neutralization of AHG reagent by residual
unbound serum globulins
Saline for washing Fresh or buffered at pH 7.2 to 7.4
Addition of AHG Immediately
Centrifugation 1000 RCF for 20 seconds
39
7
Reagent Action Procedure Antibody ID
Saline 4-22⁰C Immediate spin IgM
up to 60 min
37⁰C for 45-60 min IgG
AHG Cross-links sensitized cells, DAT a) Polyspecific: anti-IgG

resulting in visible IAT + anticomplement
agglutination b) IgG monospecific:
anti-IgG
22% Adjust zeta potential Incubation at 37⁰C for IgG

Albumin between RBCs 45-60 min
LISS Causes RBC to take up Incubation at 37⁰C for 5- IgG

antibody more rapidly 15 min
PEG Increases test sensitivity; Incubation at 37⁰C for IgG

aggregates RBC causing 10-30 min
closer proximity to assist in
Ab cross-linking
39
8
Reagent Action Procedure Antibody ID
Enzymes Reduces RBC charge; a) One-step Destroy:

Destroys or depresses b) Two-step Duffy, MNS
some RBC antigens;
enhances other RBC Enhance:
antigens Rh, Ii, Kidd, Lewis,
P1
PROTEOLYTIC ENZYMES
a) Ficin
b) Papain
c) Bromeli
d) Trypsin
39
9
Reagent Mode of action
Dithiothreitol Break the disulfide bonds of the J chain of the IgM
β-2-mercaptoethanol molecule but leave the IgM molecule intact
ZZAP Dissociation of IgG molecules from the surface of
sensitized RBCs and alters the surface antigens of
RBCs
Papain Cleaves Ig above the hinge region
3 Fragments: 2 Fab + Fc
Pepsin Cleaves Ig below the hinge region
2 Fragments: F(ab)2 + Fc’
40
0
40
1
Antibody Neutralizing Substances
Anti-P1 Hydatid cyst fluid, pigeon
droppings, turtledoves’ egg
white
Anti-Lewis Secretor saliva, plasma or
serum
Anti-Chido, Anti-Rodgers Plasma or serum (pooled)
Anti-Sda Guinea pig urine
Anti-I Mother’s milk
40
2
9 Compatibility Testing 10 Testing for antibodies
40
3
40
4
 Destruction of the RBCs of the fetus and
neonate by the antibodies produced by
the mother
 Only antibodies of the immunoglobulin G
(IgG) class are actively transported
across the placenta
40
5
40
6
40
7
40
8
1. Anemia
• Compensated anemia (Inc immature RBCs=
erythroblastosis fetalis)
2. Generalized edema and cardiac feature
(hydrops fetalis)
3. Increase urinary bilirubin
4. Deposition in brain tissue (kernicterus)
5. Hepatosplenomegaly (extramedullary
hematopoiesis)
40
9
Pathologic Characteristics
Feature
In Utero Hydrops Anemia
fetalis Cardiac insufficiency
Accumulation of fluid in the
tissues of the fetus (edema)
Neonatal Permanent, Antibody-mediated
period irreversible hemolysis
brain Accumulation of plasma
damage bilirubin
Absorption of bilirubin by
body tissue containing lipids,
including the brain
41
0
Characteristics ABO Rh
Antibody Non immune/ immune IgG Immune IgG anti-D, etc.
anti-A, B
Blood group Mother=O Mother= Rh Negative
Baby=A/ B/ AB Baby= Rh Positive
Obstetric history First pregnancy and First pregnancy not
subsequent pregnancies may affected; Rare
be affected
Disease predicted No Yes
by titers
Bilirubin at birth Normal Elevated
Anemia at birth No Yes
DAT Weakly positive/ negative Positive
Spherocytosis Yes Rare
Therapy Phototherapy Phototherapy, exhange/
intrauterine transfusion 41
1
Intrauterine Exchange
Transfusion Transfusion
Definition Transfusion of Removal of infant RBCs coated with
packed RBCs to the maternal antibody and replacement
fetus with antigen negative RBCs
When In utero Neonatal period
performed
Goals Correction of Removal of bilirubin
anemia Removal of antibody
Removal of antibody coated RBCs
Provision of antigen negative RBCs
Sample for Mother’s serum Mother’s serum (pref) or infant’s
crossmatching serum or eluate made from cord RBCs
Selection of Fresh, Group O Fresh, Group O or group specific,
RBCs negative for CMV neg, HbS neg
offending antigen,
CMV neg, HbS neg 41
2
41
3
41
4
 Shortened RBC survival mediated
through immune response, specifically
by humoral antibody
1. Alloimmune
2. Autoimmune
3. Drug-induced
41
5
1. Alloimmune
 Patient produces alloantibodies to
foreign or non-self RBC antigens
introduced into the circulation, most
often through transfusion or pregnancy.
41
6
2. Autoimmune
 Patient produces antibodies against his
or her own RBC antigens
41
7
3. Drug-induced
 Production of antibodies to a particular
drug or drug complex, with ensuing
damage to the individual’s RBCs
41
8
Characteristics WAIHA CHAD PCH
DAT + + +
Donath-Landsteiner - - +
test
Ab class IgG IgM Biphasic IgG
Optimal reaction 37C 0-4C 0-4C (antibody
temp binds to cell
surface)
37C (hemolysis)
Blood group Rh, Kell, I, I P
specificity others
Primary Extravascul Extravascular, Intravascular
mechanism of cell ar, splenic hepatic
destruction
41
9
11 HDFN
42
0
1. ABO and Rh blood group systems 5%
2. Other major blood group systems: Kell, Duffy, Kidd, Lewis, MNSs, 3%
Lutheran, P, I
3. Minor blood group systems: Diego, Cartwright, Chido, XG, Scianna, 1%
Gerbisch, Milton, Knops, Bg, Indian, etc.
4. Basic genetics 2%
5. Blood donor selection and processing 5%
6. Blood preservation and banking 5%
7. Component preparation 5%
8. Transfusion therapy 2%
9. Transfusion reactions 3%
10. Transfusion-transmitted diseases 3%
11. BB techniques and procedures: typing, compatibility testing, antibody 8%
detection and identification
12. Hemolytic disease of the newborn and autoimmune hemolytic anemia 4%
13. Quality management (structure, set-up/ equipment) 4%
42
1
IN BLOOD BANKS
42
2
 “Degree of excellence”
• Safe, satisfying donation experiences for blood
donors
• Accurately labeled and tested blood
components provided to transfusion services
• Timely, accurate transfusion services provided to
physicians and other health care personnel
• Safe and efficacious blood transfusions to
patients
42
3
Quality Management
Activities of the management function that
determine the quality policy
Quality Quality Quality

System Assurrance Control
Planned, systematic
Organizational structure,
activities implemented
procedures, processes and Operational techniques and
within the quality system to
resources needed to activities used to fulfill the
provide confidence that
implement quality requirements for quality
requirements for quality will
management
be fulfilled
42
4
Monitoring of Instruments or Equipment
Mercury thermometers Annually
Cell washers, blood warmers, centrifuge Quarterly
(speed timer)
Centrifuge (refrigerated) Monthly
Alarm activation (ref and freezers) Monthly
Blood typing reagents, heating blocks, water Daily
baths, refrigerators and freezers
(continuous), platelet incubators (enclosed,
monitored)
Platelet incubators (ambient temp) Every four
hours 42
5
Monitoring of Instruments or Equipment
Donor unit agitators Daily when in use
Scales Daily when in use
Balances Daily when in use
Hemoglobinometer Daily when in use
Microhematocrit centrifuges Daily when in use
42
6
Set of planned actions to provide
confidence that systems and
elements that influence the quality of
the product or service are working
as expected
Addresses how well an entire
process is functioning
42
7
QC Activities QA Indicators
•Collection equipment •Number of donor forms
(Microhct, Hb instrument, with incomplete or
etc.) incorrect information
•Blood components •Number and types of
•Reagents (CuSO4, antisera, unusable units and blood
etc.) components
•Laboratory equipment •Number of blood typing
(Heating instruments, water discrepancies in donors
baths, centrifuges, etc.) and patients
42
8
Collections, Components, Testing,
Distribution, Compatibility Testing,
Administration
Quality System Essentials:
Organization
Personnel
Equipment
Purchasing and Inventory
Process Control
Documents and Records
Occurrence Management
Assessments: Internal and External
Process Improvement
Facility and Safety
42
9
Policy What will be done
Process How it happens
Procedure How to do it
43
0
Plan-Do-Check-Act (PDCA)
Plan
A mission-consistent, customer-oriented action plan
Do
Put the plan into action
Check
Has the planned and implemented change created
intended improvement?
Act
Decide what to do next
43
1
• Determine product/ service specifications
Plan • Prepare SOPs
• Appraise conformance to standards

Implement • Act on difference
• Perform ongoing audits

Assess
• Train staff in use of problem-solving methods and tools

Improve • Monitor impact of process improvements
43
2
IN THE BLOOD BANK
43
3
HIS/LIS
Assistin the management of data
and can allow tracing of a blood
component through all
processing stems from donation
through transfusion
43
4
Computer system
• Hardware
• Software
• People
43
5
43
6
 Physical pieces of equipment
 Box: central system unit
• Processing hardware
 Central Processing Unit (CPU)/ processor
 ROM and RAM chips
• Input devices
 Keyboards, pointing devices, bar code readers,
testing instruments, modem
• Output devices
 Monitor, printer, modem
43
7
 Tellsthe computer what to do with all of
the information it has received
• Operating system software
 Controls the computer’s hardware, manipulates the
application software and coordinates flow of data
• Application software
 Perform specific tasks (word processing,
spreadsheets, databases)
• Interface software
 Allows the system to communicate with other
computer systems
43
8
 The human components of a blood bank
information system
 System manager and users
43
9
12 Serological Testing 13 Problem Solving
44
0
“There isn’t any map which shows the road to
success. You have to make it yourself.”
44
1
Blood Banking Review
44
2

Blood Banking Review

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Medical Technology Review

by ANDRE KARL S. FACULIN, RMT

50%=Immunology & Serology

 1628– William Harvey discovered

 1902: vonDescatello & Sturle/

 Edward Lindemann: first to use

 1908– Carlos Moreschi: Antiglobulin

 1916– Rous and Turner: citrate-dextrose

 1932– First blood bank in Leningrad,

 1943–Loutit and Mollison introduced

 1957– Gibson introduced improved

 Homozygous (identical alleles)

Figure. Pedigree Chart

• Law of independent segregation

(p+q)2= p2 (AA) + 2pq (Aa) + q2 (aa)

Mostly IgM cold agglutinins Mostly IgG warm antibodies

BLOOD REACTION WITH REACTION WITH REACTION WITH

Note: If label of anti-sera is removed, check with controls!

BLOOD GROUP REACTION WITH A CELLS REACTION WITH B CELLS

Universal donor for RBCs

Universal donor for plasma/plasma products

Universal donor for RBCs O

Universal donor for plasma/plasma products AB

H Blood Group I/i

Blood Group Antigen Present Anti-A Anti-A1 lectin

Blood Group Antigen Present Anti-A Anti-A1 lectin

ABO Group Secretor A cells B cells O cells

Anti-A Anti-B A1 cells B cells O cells Autocontrol

Anti-A Anti-B Anti-AB A1 cells B cells RBC %

0 2+mf 2+mf 4+ 0 70% B; 30% O

0 +wk +wk 4+ 0 30% B; 70% O

Px phenotype Anti-A Anti-B A1 cells B cells

Anti-A Anti-B A1 cells B cells

Anti-A Anti-B A1 cells B cells

Anti-A Anti-B A1 cells B cells O cells Autocontrol

Anti-A Anti-B A1 cells B cells O cells Autocontrol

Anti-LW is different from anti-D

Factor Rh0 Rh0

Rh0 gene Factor hr’ hr’

Factor hr’’ Hr’’

 ISBT : Numeric technology

IgM anti-D reagents Immediate spin saline testing

Chemically modified IgG Direct saline agglutination testing

Monoclonal anti-D Single clones of antibody-producing

Monoclonal blends Combinations of monoclonal IgM

Du typing (Test for Weak D)

Kleihauer-Betke test reveals 3% fetal cells. How

lele Lea- Leb-

le se Lea- Leb- Lec+

le Se Lea- Leb- Lec- Led+

During pregnancy, antigens become weaker

Le a+b- 22% 23%

Le a-b+ 72% 55%

• ENZYMES ENHANCE ACTIVITY

• ABSORPTION IS USED TO TEST FOR OTHER

Numeric Alpha Name Incidence__________

• Anti-Lub - rare - mostly IgG and requires

• Other antibodies react with rare Lu

Anti-Ch Anti-Rg Anti-Kn Anti-JMH

Anti-Yka Anti-Csa Anti-McC

Hematocrit ≥33% ≥38%

 Prime the system: NSS

 Equivalent to 6 to 10 Random Platelet

 Corticosteroids, G-CSF, GM-CSF