Exercise 4 – Blood types

Red blood cells (RBCs) have characteristic molecules on the surface of their membranes that can
be different in different people. These genetically determined membrane molecules can function
as antigens – capable of binding to specific antibodies when exposed to plasma from a person
with a different blood type. The major blood group antigens are the Rh antigen and the antigens
of the ABO system.
Objectives
1. Explain what is meant by the term blood type and identify the major blood types.
2. Explain how agglutination occurs and how agglutination tests can be used to determine a
person’s blood type.
3. Identify the different genotypes that can produce the different blood group phenotypes
and explain how different blood types can be inherited.
4. Explain how erythroblastosis fetalis is produced.
5. Explain the dangers of mismatched blood types in blood transfusions.
Materials
1. Sterile lancets
2. 70% alcohol
3. Anti-A, anti-B and anti-Rh sera
4. Slide warmer
5. Glass slides
6. Toothpicks
7. Cotton
8. Containers for the disposed blood-containing objects
When blood of one person is mixed with plasma from another person, the red blood cells will
sometimes agglutinate, or clump together (Fig. 1). This agglutination reaction, which is very
important in determining the safety of transfusions (agglutinated cells can block small blood
vessels), is due to mismatch of genetically determined blood type.
On the surface of each red blood cell are a number of molecules that have antigenic properties,
and in the plasma each antibody molecule has two combining sites for antigens. In a positive
agglutination test, the red blood cells clump together because they are combined through
antibody bridges.
A. The Rh Factor
One of the antigens on the surface of red blood cells is the Rh factor (named because it was first
discovered in rhesus monkeys). The Rh factor is found on the red blood cell membranes of
approximately 85% of the people in the United States. The presence of this antigen on the red
blood cells (an Rh positive phenotype) is inherited as a dominant trait and is produced by both
the homozygous (RR) genotypes and the heterozygous (Rr) genotype. Individuals who have the
homozygous recessive genotype (rr) do not have the antigen on their red blood cells and are said
to have the Rh negative phenotype.
Suppose an Rh positive man who is heterozygous (Rr) mates with an Rh positive woman who is
also heterozygous.

Here, since the mother is Rh positive, her immune system cannot be stimulated to produce
antibodies by the presence of an Rh positive fetus. Since the development of immunological
competence does not occur until shortly after birth, an Rh negative fetus in an Rh positive
mother would not yet have an immune response during normal gestation (pregnancy).
However, when an Rh negative mother is carrying an Rh positive fetus, some of the Rh antigens
may enter her circulation when the placenta tears at birth (and blood cells do not normally cross
the placenta during pregnancy). Since these red blood cells express an antigen (the Rh factor)
that is foreign to the mother, her immune system will eventually be stimulated to produce
antibodies that are capable of destructing the red blood cells of subsequent Rh positive fetus, a
condition known as Hemolytic disease of the newborn or erythroblastosis fetalis. However,
erythroblastosis fetalis can be prevented by the administration of exogenous Rh antigen known
as Rho(D) immune globulin (e.g. RhoGAM) to the mother within 72 hours after delivery. These
antibodies destroy the fetal Rh positive red blood cells that have entered the maternal circulation
before they can stimulate an immune response in the mother.

(A) (B)
Fig. 1. An agglutination reaction. (A) Type A red blood cells are mixed with anti-A antibodies. (B) The results in the
formation of antigen-antibody bridges that cause the red blood cells to clump together (agglutinate).

Procedure
1. Place one drop of anti-Rh serum on a clean glass slide.
2. Add an equal amount of fingertip blood and mix it with the antiserum (use an applicator
stick or a toothpick).
3. Place the slide on a slide warmer (45oC to 50oC) and rock it back and forth.
4. Examine the slide for agglutination. If no agglutination is observed after a 2-minute
period, examine the slide under the low power objective of the microscope. The presence
of grains of agglutinated blood cells indicates Rh positive blood.
Caution: Handle only your own blood and be sure to discard the slide, toothpick, and lancet in
the container provided by the instructor.
B. The ABO antigen system
Each individual inherits two genes, one from each parent, that control the synthesis of red blood
cell antigens of the ABO classification. Each gene contains the information for one of three
possible phenotypes: antigen A, antigen B, or no antigen (written O). Thus, an individual may
have one of six possible genotypes: AA, AO, BB, BO, AB, or OO.
An individual who has the genotype AO will produce type A antigens just like an individual who
has the genotype AA; and therefore both are said to have type A blood. Likewise, an individual
with the genotype BO and one with the genotype BB will both have type B blood. Since lack of
antigen is a recessive trait, an individual with type O blood must have the genotype OO.
Unlike many other traits, the heterozygous genotype AB has a phenotype that is different from
either of the homozygous genotypes (AA or BB). Since there is no dominance between A and B,
individuals with genotype AB produce red blood cells with both the A and B antigens (a
condition known as codominance) and have type AB blood. The most common blood types are
type O and type A, the rarest is type AB (Table 1).
Also, unlike the other immune responses considered, antibodies against A and B antigens are not
induced by prior exposure to these blood types. A person with type A blood, for example, has
antibodies in the plasma against type B blood even though that person may never have been
exposed to this antigen. A transfusion with type B blood into the type A person would be
extremely dangerous because the anti-B antibodies in the recipient’s plasma would agglutinate
the red blood cells in the donor’s blood. The outcome would be the same if the donor with type
A and the recipient would be type B (Fig. 1b).
Procedure
1. Draw a line down the center of a clean glass slide with a marking pencil and label one
side A and the other side B.
2. Place a drop of anti-A serum on the side marked A and a drop of anti-B serum on the side
marked B.
3. Add a drop of blood to each antiserum and mix each with a separate applicator stick.
4. Tilt the slide back and forth and examine for agglutination over a 2-minute period. Do not
heat the slide on slide warmer.
5. Enter your ABO blood type in the laboratory report.
 HUMAN PHYSIOLOGY AND PATHOPHYSIOLOGY

Name: _________________________________ Date: ______________

Section: ________________________________

Exercise 4 – Blood types

Data:
1. Did your blood agglutinate with the anti-Rh serum? _____________________________
Are you Rh positive or negative? ____________________________________________
2. Indicate below (with a yes or no) whether your blood agglutinated with the anti-A and
anti-B sera.
Anti-A: __________
Anti-B: __________
What is your blood type? ______________

Questions:
1. Name the antigens present and absent on the surface of a person’s red blood cells if that
person is:
(a) Type A negative __________
(b) Type O positive ___________
(c) Type AB negative _________
2. What are the dangers of giving a person with type A negative blood a transfusion of type
B positive blood?

3. Explain how hemolytic disease of the newborn is produced. How may this disease be
prevented?

4. In a paternity suit, a woman (type O) claims that a man (type A) is the father of her baby
(type O). Will the blood types prove or disprove her claim? Explain.