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Hematology
What is Karl Landsteiner credited with
discovering?
Blood
Composition
Key Terms
• Anticoagulant: an agent that prevents the
clotting of blood.
– Examples are EDTA, Citrate and Heparin
• Capillary: small blood vessel that connects
arterioles and venules
• Hematoma: a subcutaneous mass of blood
at a venipuncture site
Key Terms
• Hemoglobin: the oxygen carrying molecule
of red blood cells
• Hemolysis: the breakdown of red blood
cells, with the release of hemoglobin into
the plasma or serum. Cannot use
hemolyzed samples in lab tests
• Icteric: jaundiced; dark yellow or greenish
serum or plasma
Key Terms
• Lipemic: having abnormally high level of
fat. Milky looking samples
• Plasma: pale yellow part of whole blood;
contains all clotting factors
• Serum: liquid portion of blood without the
protein fibrinogen, which is one of the
clotting factors; clot removed
Key Terms
• Agglutination: the clumping together of
rbc by the action of an antibody
– When A cells are added to a blood sample,
agglutination will occur only in the presence of
anti-A
• Serology: the study of antigen – antibody
reactions using laboratory tests
Function of Blood
• Transporting fluids such as:
–
–
–
–
Nutrients from digestive tract
O2 from lungs
Waste from cells
Hormones
• Aids in heat distribution
• Regulates acid-base balance
Composition of Blood
• Plasma: liquid portion of blood w/out cells
– Contains all of the following
• Water
• Electrolytes
Nutrients
Metabolic waste
product
• Hormones
Vitamins and enzymes
• Plasma proteins such as fibrinogen, albumin and
globulin
Composition of Blood:
Erythrocytes
• Red blood cells are responsible for:
– Transport of oxygen and nutrients
– Removal of waste and CO2 from the cells
– Distribution of heat
• Hemoglobin: the O2 carrying potential
Composition of Blood:
Leukocytes
• WBC are responsible for:
– Phagocytosis – to engulf and absorb waste
material and harmful microorganisms in the
blood stream and tissues
– Synthesis of antibody molecules
– Inflammation process
– Production of heparin – component found in
lung and liver tissue which have the ability to
prevent clotting of blood.
• Heparin used in the treatment of thrombosis
Cell Morphology
Lymphocyte
rbc
Platelet
Segmented Neutrophil
Composition of Blood:
Leukocytes
• Types of Leukocytes
– Granulocytes
• Neutrophils
• Eosinophils
• Basophils
– Agranulocytes
• Lymphocytes
• Monocytes
Composition of Blood:
Thrombocytes
• Platelets – the smallest of the solid
components of the blood
• Responsible for the clotting process
• Coagulation: term for clotting
• Embolism: a blood clot which is moving
through the body
Forensic Characterization of
Blood
• 1st questions a criminalist has to answer is:
– Is it blood
– If yes, is it human
– If yes, can it be associated w/ a particular person
• Preliminary color test for blood is the
Benzidine color test but this is carcinogenic
so phenolphthalein is used and is known as
the Kastle-Meyer Color test
Forensic Characterization of
Blood
• Hemoglobin possesses peroxidase like
activity which when mixed with
phenolphthalein and hydrogen peroxide it
will cause the formation of a deep pink
color
• Kastle-Meyer is not specific for blood as
some vegetables such as potatoes and
horseradish contain peroxidase and can
react; however they should not be
common at a crime scene so it is often
considered a good indicator
Forensic Characterization of
Blood
• Hemastix strips can also be used to detect
the presence of blood
• Luminol test is another presumptive test
for blood which produces light rather than
a color reaction. Objects being tested
must be in a dark location to view the
luminescence (emission of light).
– Extremely sensitive and can detect blood
diluted up to 300,000 times
– Luminol will not interfere with other DNA
testing
Forensic Characterization of
Blood
• Once blood is found it must be determined
to be human
– Precipitin test is the standard test
– Reagents are available to determine if blood is
dog, cat or deer
– A positive test is a cloudy ring or band at the
point where the two liquids meet
– Only a small amount of sample is required
– Precipitin test is very sensitive and can test
positive on a sample as old as 10 years
– Extracted tissue samples from mummies as old
as 4,000 years have tested positive
Forensic Characterization of
Blood
• Gel diffusion is another method which can
be used.
• Antigen and antibody assay
• Gel electrophoresis can also be used to
determine if a sample is human blood or
not
• Antigen and antibody assay
• Uses electrical current and ppt. line forms where the
two samples meet if it is a positive reaction
Blood Types
• Four Major Groups
–A
B
AB
O
• Blood types are inherited from your
parents
• Antigen is present on the red blood cell;
typing is done w/rbc
• Antibody is present in the plasma;
antibody screening done on plasma
Blood Types
• O negative
– Universal donor
– It carries no antigen
• AB positive
– Universal recipient
– It carries no antibodies in the plasma
• 43% of population are O, 42% A, 12% B
and 3% AB
Rh Factor (D antigen): found
on the surface of rbc
• Rhesus factor:
discovered in rhesus
monkeys in 1937
• Can be phenotypically
positive or negative
• Positive is dominant
over negative
– If positive is present,
then you will express
positive phenotype
+
-
_
+
-
++
+-
+-
--
_
+
+- +-
+
+- +-
Blood Types: ABO
• Controlled by
GENETICS!!!!!
• Two Categories
A
O
B
AB
BO
O
AO
OO
– ABO and Rh
A
A
A
O
O
AA AO
AO OO
B
A
AA
AB
B
AB
BB
Blood Types: What Ag do
they have?
• A positive blood has which Ag present on
rbc?
– A antigen and Rh antigen
• B negative blood has which Ag present on
rbc?
– B antigen
• AB + blood type has which Ag present?
– Both A and B Ag and Rh antigen on the rbc
Who can donate to whom?
• A + donor: what blood types can this pt.
receive?
– A+, A-, O+, O-
• Your blood type is B-, what blood types
can donate to you?
– B- and O-
AB- as a last resort
ABO Blood Types
Agglutination Reactions
Anti-A
Anti- B
Anti-A &
Anti B
A
Yes
No
Yes
B
No
Yes
Yes
AB
Yes
Yes
Yes
O
No
No
No
Examples of ABO blood typing
O negative carries no Ag and therefore does not react with any Anti
A, B, AB
Pos reaction
Neg reaction
Rh Antigen and Antibody Interaction
Anti- Rh
Rh +
YES
Rh -
NO
Normal Ranges
• RBC:
female 3.6-5.0x106mm3
male 4.2-5.4x106mm3
• WBC:
4.5-10.5x103mm3 (African Americans is sltly lower 3.2
is still normal)
• HCT:
female 36-48%
• Hgb:
female 12-16 g/dL male 14-17.4 g/dL
male 42-52%
• Platelets: 140-400x103mm3
Blood Cell
Maturation
Blood Splatter Analysis
• Location, distribution, and appearance of
blood stains are an important part of
forensics
• Investigators try to determine:
– Direction
– Dropping distance
– Angle of impact
• Splatter analysis is often used for crime
scene reconstruction
Blood Splatter Analysis
• Factors which influence stain patterns are:
– Surface texture
– Direction of travel
• Pointed end of bloodstain always faces its direction
of travel
– Angle of impact is determined by measuring
the degree of circular distortion of the stain
• Blood striking a surface at right angles gives rise to a
nearly circular stain
• As the angle decreases, the stain becomes
elongated in shape
DNA
•
•
•
•
•
•
DeoxyriboNucleic Acid
Found in the nucleus
46 chromosomes
25,000 genes
Structure determined by Crick and Watson
DNA fingerprinting by Alec Jeffreys
Structure of DNA
• A polymer made of repeating nucleotides
• Nucleotide consists of a deoxyribose sugar,
a phosphate, and a nitrogen base
(Adenine, Thymine, Cytosine, or Guanine)
• Double stranded, helical
• Complementary base
pairing, A=T, G=C
DNA at Work
• DNA in nucleus is copied into a strand of
RNA (transcription)
• RNA is read at the ribosome to make
assemble amino acids into proteins
(translation)
• Every 3 bases on DNA
codes for a different
amino acid
Replication of DNA
• Replication – the synthesis of new DNA
from existing DNA in the nucleus
• DNA polymerase assembles
new DNA strand and proofreads it
• Replication occurs in nucleus prior to cell
division
Polymerase Chain Reaction
• A technique for replication, or amplifying,
a portion of DNA outside
the cell
• Each cycle doubles
the number of copies
• 1
1x107 in 30 cycles
DNA Typing with Tandem
Repeats
• Region of chromosome that contains multiple copies of a
core DNA sequence arranging in a repeating fashion
between the coding regions (genes)
• Restriction Fragment Length Polymorphisms used
enzymes to cut the DNA around these tandem repeat
sites and then run them on a gel electrophoresis
• A Southern blot was then performed and radioactive
probes were hybridized to help visualize the RFLPs
RFLPs
PCR
PCR has the following advantages:
1. PCR can use shorter sequences
2. shorter pieces more stable
3. smaller amounts of DNA can
be used (10-9 gram)
Short Tandem Repeats
(STRs)
• A region of a DNA molecule that contains
short segments of 3-7 repeating base
pairs.
• Generally less than 450 bp long
• Less degradation
• Can be PCR’d
• Can multiplex a large number of these
STRs at once
• US uses 13 STRs for tests
• Capillary electrophoresis
• Sex Identification by focusing on the
amelogenin gene