M11A: ABO and RH Blood Typing Discussion
then the antigen is present.
ABO and RH Blood Typing
o Is one of most important tests in the clinical laboratory in
particularly, in the blood bank
o It helps us to identify the specific antigens present on the
surface of the of the red blood cells which determines the
patient’s blood type.
Blood Typing
o Determine the antigenic substances present on the
surface of the red blood cells (These antigenic substances
could elicit a strong immune response)
o Blood typing is a medical laboratory test used to determine
the antigen present on the external surface of the Red
blood cells (RBCs)  to determine  a person’s blood type.  3. With a clean glass slide, collect three separate drops of blood.
o Routine blood typing includes the determination of the
most immunogenic (may produce a strong immune response
such as antibody production) antigen in humans, The
ABO and Rh.
ABO Blood Group System / Blood Classification
o There are over 40 blood group system already discovered,
but the most immunogenic blood group system is the “ABO
blood group system”— On this blood group we could
identify the a and b antigen. The ABO blood type
classification was discovered by Austrian scientist named
Karl Lansdsteiner in 1901. This blood group system is used
to indicate the presence of one, both or neither of the A and
B antigens on the surface of the RBCs. ABO blood type
system is the most immunogenic among other blood group
system. A mismatch in this blood type can cause a
potential adverse reaction after transfusion which can
lead to death.
RH blood group system
o The next immunogenic blood group system is the Rh blood
group system—we identify the d antigen.
o The RH blood group system is the second most important
blood group which was discovered in 1937 by Karl
Landsteiner and Alexander S. Wiener. The term “Rh” was
due to the fact that it was first elucidated on the RBCs of a
rhesus monkey. There are over 50 antigens on these blood
group systems but D, C, E, c and e antigen are the most
important. This blood group system will denote if a person
is Rh positive or Rh negative. Blood typing could either be
performed using slide method, tube method and gel
method. For this activity we will perform the simplest
method which is the slide blood typing.
Forward slide typing
o Involves the use of known antibody to identify the
unknown antigen.
Blood typing together with cross matching blood typing is a significant
test prior to blood transfusion to ensure its safety.
Principle of Blood Typing
o If the antigen is present on the blood, the homologous
antibody present on the antisera reacts to it, causing
agglutination.
o Will use anti-a, anti-b and anti-d antisera.
o If the blood has agglutination on that specific antisera,
o But if there is no agglutination on the specific antisera,
then the antigen is not present.
ABO AND RH BLOOD TYPING PROCEDURE
Materials: alcohol swab, slide applicator stick, lancet, anti-a
antisera (blue), anti-b antisera (yellow), and anti D antisera
(clear)
1. Using cotton with alcohol, sterilize the finger to be pricked
and wipe the area dry with another cotton.
2. Pricked the finger using a blood lancet and wipe the first
drop of blood to avoid contamination of the tissue juice or fluid.
4. Immediately add a drop of Anti-A reagent to the first drop of
blood and gently mix it using applicator stick.
5. Subsequently, add a drop of Anti-B reagent to the second
drop of blood and mix using a new applicator stick.
6. Lastly, add a drop of Anti-D reagent to the third drop of
blood and mix using a new applicator stick.
7. Observe the formation of visible agglutination.
Agglutination = is a reaction in which particles such as red
blood cells suspended in a liquid, form a clump and which
occurs specially as a serologic response to a specific antibody.
M11B: Clotting Time and Bleeding Time Discussion
Clotting time and Bleeding Time
o Clotting time and Bleeding time are routine tests
done to evaluate the coagulation and hemostasis.
o Clotting time is the measure of rate of fibrin thread
formation in vitro. 
o While Bleeding time is a test wherein the rate for a
wound to stop bleeding is measured through the
platelet-plug formation. 
o Both of these tests are considered as an important
pre-surgical procedure to ensure the safety of the
operation as it helps in identifying bleeding
tendencies.
Hemostasis
o is a hematological process to arrest and stop the
bleeding. It involves series of reactions designed to
maintain the blood inside the blood vessels whenever
the tissues are damaged.
Coagulation
o is the process where blood is transformed from liquid
to gel.
Clotting Time
o is a measure of coagulation wherein the time for a
blood sample to coagulate in vitro is measured
o Time required for blood form a clot (a clot is a solid
end product of a coagulation pathway)
o Glass slide initiated the intrinsic pathway of
coagulation to form a clot (We’ll stop and will take
note of the time, once we observed the formation of
fibrin thread—indicates that the blood has already
be clotted)
o Principle: A blood sample is placed in a slide to
measure the ability of the clotting factors to form a
clot. The time required for the blood to form a fibrin
thread is recorded and termed as the clotting time.
o There are various methods for determining the
clotting time, the most commonly used is the
capillary tube method. However, for this activity we
will perform the micromethod or slide method of
clotting time, this method is most commonly used in
 clinical settings because it is faster and easier to
perform while in the bedside of the patient.
o Normal value of clotting time is 2-7 minutes.

Bleeding Time
o is a measure of hemostasis and coagulation. It
depends on various factors particularly on the
function of platelets and endothelial cells of arteries. 
o How quickly the blood clots to stop bleeding
o The cessation of bleeding indicates the formation
of platelet plug.
o Therefore, we can say that this test depends on the
adequacy and quality of platelets.
(inadequate/low=longer BT; adequate/good=shorter)
o Principle: The time how long the incision of the
fingertip bleeds is measured. Cessation of bleeding
indicates the formation of haemostatic plug and it is
termed as the bleeding time.
o This test measures how long it takes for the blood to
stop the bleeding. This is usually performed by
making a small incision on the skin. For this
exercise, we will use the Duke’s method of bleeding
time determination.
o Usually, the normal value of bleeding time is between
2-5 minutes.
o We could also derived the value of the bleeding time
by multiplying the blood (spot/clot?) of the filter
paper to 30 seconds.

Materials: Glass slide, lancet, alcohol swab, cotton, filter paper,

timer
CLOTTING TIME PROCEDURE
1. Using cotton with alcohol, sterilize the finger to be pricked.
2. Prick the finger using a blood lancet and collect three dome-
shaped blood drops on the slide. Start the timer once the blood
started to ooze out from the finger.
3. Using a lancet or a needle, check if there is a clot or fibrin-
thread formation by drawing the blood every 30 seconds.
4. Take note of the clotting time once the formation of fibrin
clot was observed. (2 min and 30 seconds)

BLEEDING TIME PROCEDURE

1. Sterilize the finger to be pricked using cotton with alcohol.
2. Prick the finger with blood lancet and record the time when
blood starts to ooze from the finger. (Avoid applying pressure
to the pricked finger)
3. Blot the blood every 30 seconds using the filter paper.
4. Once the bleeding ceases, stop the timer and record the
bleeding time. (1 min and 30 seconds)