INTRODUCTION to

IMMUNOHEMATOLOGY
BLOOD BANKING & TRANSFUSION MEDICINE

BLOOD BANKING encompasses activities, procedures, and tests

done to ensure that blood for transfusion is properly collected,
preserved, stored, and dispensed for later use in blood transfusion.

TRANSFUSION MEDICINE is a multidisciplinary branch of

medicine concerned with the transfusion of blood and blood
components, including proper selection and utilization of blood
components as well as removal of blood or blood components in
the treatment or prevention of disease.
 History & Trends of the
Blood Transfusion Practice
 HISTORY OF TRANSFUSION MEDICINE

▪ 1492 First recorded unsuccessful blood transfusion;

donors were three young men in the hope of curing the
stricken Pope Innocent VII
▪ 1628 English physician William Harvey discovered the
circulation of blood.
▪ 1665 The first recorded successful blood transfusion
occurs in England: Physician Richard Lower kept dogs
alive by transfusion of blood from other dogs.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1818 James Blundell, a British obstetrician, performs

the first successful transfusion of human blood to a
patient for the treatment of postpartum hemorrhage.
▪ 1840 At St. George's School in London, Samuel
Armstrong Lane, aided by consultant Dr. Blundell,
performs the first successful whole blood transfusion to
treat hemophilia.
▪ 1867 English surgeon Joseph Lister used antiseptics to
control infection during transfusions.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1869 Braxton Hicks recommended sodium phosphate

in attempt to find a nontoxic anticoagulant as clotting
was the principle to overcome.
▪ 1873-1880 US physicians transfuse milk (from cows,
goats, and humans).
▪ 1884 Saline infusion replaces milk as a “blood
substitute” due to the increased frequency of adverse
reactions to milk.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1901 Karl Landsteiner, an Austrian physician,

discovered the ABO blood groups. He received the Nobel
Prize for Medicine for this discovery in 1930.

▪ 1902 His colleagues Alfred Decastello and Adriano

Sturli add AB, the fourth blood group.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1907 Hektoen suggests that the safety of transfusion

might be improved by crossmatching blood between
donors and patients to exclude incompatible mixtures.
Reuben Ottenberg performs the first blood transfusion
with performed blood typing and crossmatching, in New
York. Ottenberg also observed the mendelian inheritance
of blood groups and recognized the “universal” utility of
group O donors.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1908 French surgeon Alexis Carrel devised a way to

prevent clotting by sewing the vein of the recipient
directly to the artery of the donor. This vein-to-vein or
direct method, known as anastomosis, was practiced by a
number of physicians, among them J.B. Murphy in
Chicago and George Crile in Cleveland. The procedure
was proved unfeasible for blood transfusions, but paved
the way for successful organ transplantation, for which
Carrel received the Nobel Prize in 1912.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1908 Moreschi describes the antiglobulin reaction. The

antiglobulin is a direct way of visualizing an antigen-
antibody reaction that has taken place but is not directly
visible. The antigen and antibody react with each other,
then, after washing to remove any unbound antibody, the
antiglobulin reagent is added and binds between the
antibody molecules that are stuck onto the antigen. This
makes the complex big enough to see.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1912 Roger Lee, a visiting physician at the

Massachusetts General Hospital, along with Paul Dudley
White, develops the Lee-White clotting time. Adding
another important discovery to the growing body of
knowledge of transfusion medicine, Lee demonstrates
that it is safe to give group O blood to patients of any
blood group, and that blood from all groups can be given
to group AB patients. The terms "universal donor“ and
"universal recipient“ are coined.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1913
Vein-to-vein
blood transfusion
was carried out by
Edward Lindemann
using multiple
syringes and a special
cannula for
puncturing the vein
through the skin.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1914 Hustin reported the use of sodium citrate as an

anticoagulant, allowing longer preservation of blood.

▪ 1915 Richard Lewisohn used sodium citrate as an

anticoagulant to transform the transfusion procedure
from direct to indirect.
▪ In addition, Richard Weil demonstrated the feasibility of
refrigerated storage of such anticoagulated blood.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1916 Francis Rous and J.R. Turner introduced a citrate-

glucose solution that permits storage of blood for several
days after collection.

▪ Oswald Robertson, an American Army officer, was

credited with creating the blood depots. He received the
AABB Landsteiner Award in 1958 as developer of the first
blood bank.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1927-1947 The MNSs and P systems are discovered.

▪ 1939/40 The Rh blood group system was discovered by

Karl Landsteiner, Alex Wiener, Philip Levine, and R.E.
Stetson and was soon recognized as the cause of the
majority of transfusion reactions.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1940 The United States government established a nationwide

program for the collection of blood. Charles R. Drew developed
the “Plasma for Britain” program — a pilot project to collect blood
for shipment to the British Isles. The American Red Cross
participated, collecting 13 million units of blood by the end of
World War II.
▪ 1941 Dr. Drew was appointed director of the first American Red
Cross Blood Bank at Presbyterian Hospital.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1943 The introduction by J.F. Loutit and Patrick L. Mollison of

acid citrate dextrose (ACD) solution

▪ 1943 P. Beeson published the classic description of transfusion-

transmitted hepatitis.

▪ 1945 Coombs, Mourant, and Race described the use of

antihuman globulin (later known as the “Coombs Test”) to identify
“incomplete” antibodies.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1947 The American Association of Blood Banks (AABB)

was formed to promote common goals among blood
banking practitioners and the blood donating public.

▪ 1949-1950 The US blood collection system include 1,500

hospital blood banks, 46 community blood centers, and
31 American Red Cross regional blood centers.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1950 Audrey Smith reported the use of glycerol

cryoprotectant for freezing red blood cells.

▪ 1950 Carl Walter and W.P. Murphy, Jr., introduced the

plastic bag for blood collection.
▪ Development of the refrigerated centrifuge in 1953
further expedited blood component therapy.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1957 The AABB formed its committee on Inspection and

Accreditation to monitor the implementation of standards
for blood banking.
▪ 1958 The AABB published its first edition of Standards
for a Blood Transfusion Service (now titled Standards for
Blood Banks and Transfusion Services).
▪ 1960 The AABB begins publication of TRANSFUSION, the
first American journal wholly devoted to the science of
blood banking and transfusion technology.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1957 Gibson introduced an improved preservative

solution called CPD – a less acidic anticoagulant.
▪ 1959 Max Perutz of Cambridge University deciphered
the molecular structure of hemoglobin, the molecule that
transports oxygen and gives red blood cells their color.
▪ 1960 A. Solomon and J.L. Fahey reported the first
therapeutic plasmapheresis procedure — a procedure
that separates whole blood into plasma and red blood
cells.
 HISTORY OF TRANSFUSION MEDICINE

▪ 1979 A new anticoagulant preservative, CPDA-1, extends

the shelf life of whole blood and red blood cells to 35
days, increasing the blood supply and facilitating resource
sharing among blood banks.

▪ 1983 Additive solutions extend the shelf life of red

blood cells to 42 days.
 CURRENT STATUS?
• Traditionally, the amount of whole blood in a unit has been 450
mL ± 10% of blood. More recently, 500 mL ± 10% of blood are
being collected.
• Units of the whole blood collected can be separated into three
components: packed RBCs, platelets, and plasma.
• In recent years, less whole blood has been used to prepare
platelets with the increased utilization of apheresis platelets.
• Whole blood-prepared RBCs may be stored for 21 to 42 days.
• Donated blood is free but a fee is still charged for each unit to
cover the costs associated with collection, storage, testing, and
transfusion.
CURRENT STATUS?
 RED BLOOD CELL:
BIOLOGY & PRESERVATION
Three areas of RBC biology are crucial for normal
erythrocyte survival and function:

❖ Normal chemical composition and structure of

the RBC membrane
❖ Hemoglobin structure and function
❖ RBC metabolism
 RBC MEMBRANE
- semi-permeable lipid bilayer supported by a mesh-like
protein cytoskeleton structure
- biochemical composition: 52% protein, 40% lipid, &
8% carbohydrate.
❖ INTEGRAL MEMBRANE PROTEINS – extend from the outer
surface and span the entire membrane of the inner cytoplasmic
side of the RBC
❖ PERIPHERAL PROTEINS – located and limited to the
cytoplasmic surface of the membrane; form the RBC
cytoskeleton
 RBC MEMBRANE
The normal chemical composition, structural
arrangement & molecular interactions of the
erythrocyte membrane are crucial to the normal
length of RBC survival in the circulation & in
RBC’s two important characteristics:
▪ Deformability
▪ Permeability
 RBC MEMBRANE
DEFORMABILITY
▪ To remain viable, normal RBCs must also remain flexible,
deformable, and permeable.
▪ The loss of adenosine triphosphate (ATP) (energy) levels
leads to a decrease in the phosphorylation of SPECTRIN
and, in turn, a loss of membrane deformability.
▪ An accumulation or increase in deposition of membrane
calcium also results, causing an increase in membrane
rigidity and loss of pliability.
 RBC MEMBRANE
PERMEABILITY
▪ The permeability properties of the RBC membrane and the
active RBC cation transport prevent colloid hemolysis and
control the volume of the RBC.
▪ Any abnormality that increases permeability or alters cationic
transport may decrease RBC survival.
▪ When RBCs are ATP-depleted, Ca2+ & Na+ accumulate
intracellularly, and K+ & H2O are lost. This results to
dehydrated rigid cell subsequently sequestered by the spleen.
 HEMOGLOBIN
▪ Primary function is gas transport: O2 delivery to tissues
& CO2 excretion
▪ Molecule is composed of four subunits, each containing
heme & globin
1 heme = 1 mole O2
1 Hb molecule = 4 moles O2
▪ One of the most important controls of hemoglobin’s
affinity for oxygen is 2,3-DPG.
 OXYGEN DISSOCIATION CURVE
▪ Graphically describes
the relationship between
oxygen content of Hb
(% of saturation) &
partial pressure of O2
(PO2)
▪ Normal shape of the
curve is SIGMOID
 OXYGEN DISSOCIATION CURVE
SHIFT TO THE LEFT
- increased Hb affinity for O2, decreased delivery of O2 to
tissues
- Increased pH, decreased 2,3-DPG, CO2, & temperature

SHIFT TO THE RIGHT

- decreased Hb affinity for O2, increased delivery of O2 to
tissues
- Decreased pH, increased 2,3-DPG, CO2, & temperature
 RBC METABOLIC PATHWAYS
Anaerobic Glycolytic Pathway
❑ Embden-Meyerhof Pathway

Ancillary Pathways
❑ Pentose Phosphate Pathway / HMP
❑ Methemoglobin Reductase Pathway
❑ Leubering-Rapoport Shunt
Blood Collection
• It is a closed system
▫ consisting of main bag with
needle, tubing, and up to four
satellite bags attached.
▫ The entire system is sterile.

• Standard phlebotomy
= 450 mL ± 45 mL (with
63 mL anticoagulant) or
= 500 mL ± 50 mL (with
70 mL anticoagulant)
 BLOOD PRESERVATION
(Anticoagulant/Preservatives)
✓ Adenine: Used in ATP synthesis
✓ Citrate: Binds calcium to prevent coagulation
✓ Dextrose: Food for the cells
✓ Phosphate: Source of 2,3-DPG which promotes oxygen release
to tissues
❑ ACD: Acid-Citrate-Dextrose (21 days)
❑ CPD: Citrate-Phosphate-Dextrose (21 days)
❑ CP2D: Citrate-Phosphate-Double Dextrose (21 days)
❑ CPDA-1: Citrate-Phosphate-Dextrose-Adenine (35 days)
 LESIONS OF STORAGE
• Biochemical changes in stored blood that can lead to
decreased RBC viability

• Glucose, ATP, 2,3-DPG, pH, and plasma sodium decrease as

RBCs are stored. After cells are transfused, ATP and 2,3-DPG levels
are restored in about 24 hours.

• Substances that increase during storage include plasma

hemoglobin, plasma potassium, ammonium, and lactic acid.
 Additive Solutions
• These are added to RBCs after removal of plasma with or
without platelets.

• They contain MAGS (Mannitol, Adenine, Glucose, Saline).

• These must be added within 72 hours of collection.

• Additives extend the shelf life to 42 days and reduce RBC

viscosity during transfusion.
 Freezing of RBCs
▪ Primarily used for autologous units and the storage of rare blood
types

Autologous transfusion (auto meaning “self”) allows individuals to

donate blood for their own use in meeting their needs for blood
transfusion
 Freezing of RBCs
ADVANTAGES DISADVANTAGES
• Long-term storage (10 years) • A time-consuming process
• Maintenance of RBC viability • Higher cost of equipment
& function and materials
• Low residual leukocytes & • Storage requirements (–65°C)
platelets • Higher cost of product
• Removal of significant
amounts of plasma proteins
 Rejuvenation Solutions
• These are used to restore ATP and 2,3-DPG levels, before
freezing or transfusing a unit, and may be necessary for
autologous or rare units.

• They contain PIGPA (Phosphate, Inosine, Glucose, Pyruvate,

Adenine).

• RBCs can be rejuvenated up to 3 days past the expiration date

(stored up to 24 hours at 1-6°C, and transfused) and can then be
frozen for future use.
 Current Trends in RBC Preservation Research
✓ Development of improved additive solutions
✓ Development of procedures to reduce and inactivate the
level of pathogens that may be in RBC units
✓ Development of procedures to convert A-, B-, and Ab
type RBCs to O-type RBCs
✓ Development of methods to produce RBCs through
bioengineering (blood pharming)
✓ Development of RBC substitutes
Current Trends in Platelet Preservation Research
✓ Development of methods that would allow platelets to
be stored for 7 days
✓ Development of additive solutions, also termed synthetic
media
✓ Development of procedures to reduce and inactivate the
level of pathogens that may be in platelet units
✓ Development of platelet substitutes
✓ New approaches for storage of platelets at 1°C to 6°C
✓ The development of processes to cryopreserve platelets
THE BASICS OF
GENETICS
 GENETICS
▪ The science of genetics is one of the most important
areas of modern biology.

▪ The understanding of the inheritance of blood group

antigens and the testing for disease markers at the
molecular level, both of which are vitally important in
transfusion medicine, are based on the science of
genetics.
 MENDEL & LAWS OF INHERITANCE
Once upon a time (1860's), in an Austrian monastery, there lived a
monk named Gregor Mendel who spent his spare time breeding
pea plants.

He did this over & over & over again, and noticed patterns to the
inheritance of traits, from one set of pea plants to the next. By
carefully analyzing his pea plant numbers, he discovered three laws
of inheritance:
1. Law of Dominance
2. Law of Segregation
3. Law of Independent Assortment
▪ Normal humans have 23 pairs of chromosomes: 22 pairs of
autosomes and 1 pair of sex chromosomes.
▪ Chromosomes are double strands of DNA that carry the genetic
information.
▪ Genes are units that code for various expressions of inherited
genetic information.
▪ Loci are specific places along the chromosomes where genes are
found.
▪ For each locus, there may be several different forms of a gene,
which are known as alleles.
▪ Blood groups are inherited in Mendelian fashion.

▪ When both the inherited alleles are identical, the person

is homozygous. If the inherited genes differ, the person is
heterozygous.

▪ Sometimes, homozygous inheritance produces a

stronger expression of the gene than would be seen in a
heterozygous individual.
The terms DOMINANT and RECESSIVE describe the inheritance
patterns of certain traits. That is, they describe how likely it is for
a certain phenotype to pass from parent to offspring.

▪ A dominant allele produces a dominant phenotype in

individuals who have one copy of the allele, which can come from
just one parent.

▪ For a recessive allele to produce a recessive phenotype, the

individual must have two copies, one from each parent.

▪ An individual with one dominant and one recessive allele for a

gene will have the dominant phenotype.
▪ Character: heritable feature (i.e., color)
▪ Trait: variant for a character (i.e. brown)
▪ True-bred: all offspring of same variety
▪ Hybridization: crossing of 2 different
true-breds

We label the different generations of a

cross as:
✓ P generation (parents)
✓ F1 generation (1st filial generation)
✓ F2 generation (2nd filial generation)
MENDEL & LAWS OF INHERITANCE
Law of Dominance
In a cross of parents that are pure for contrasting traits,
only one form of the trait will appear in the next
generation.

Offspring that are hybrid for a trait will have only the
dominant trait in the phenotype.
MENDEL & LAWS OF INHERITANCE
Law of Segregation
During the formation of gametes (eggs or sperm), the two
alleles (hereditary units) responsible for a trait separate
from each other.

Alleles for a trait are then "recombined" at fertilization,

producing the genotype for the traits of the offspring.
MENDEL & LAWS OF INHERITANCE
Law of Independent Assortment
Alleles for different traits are distributed to sex cells
(& offspring) independently of one another.
 INHERITANCE PATTERNS
Autosomal: Genes expressed with equal frequency in males and
females, on non-sex chromosome
Sex-linked dominant: Carried on the X chromosome; no father-to-
son transmission; will be expressed if passed from father to
daughter or from mother to son
Sex-linked recessive: It is carried on the X chromosome. Males
inherit it from carrier mothers; traits are exhibited most commonly
in males (e.g., hemophilia A). Females can exhibit the trait but must
inherit it from both carrier mother and affected father.