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Serologi Forensik

Farmasi Forensik

Objective
Mahasiswa akan mengetahui :

Bahwa antibodi dan


antigen dari tipe yang
berbeda akan
beraglutinasi atau
menggumpal, jika
disatukan
Bahwa signifikansi barang
bukti bergantung pada
karakteristik populasi
Peranan pengetahuan
serologi dalam ilmu
2
forensik

Objektif
Mahasiswa mampu untuk :

Menentukan apakah suatu


noda adalah darah
Menentukan tipe darah melalui
sistem ABO/Rh

Serology
Serologi adalah pemeriksaan dan analisis cairan tubuh.
Seorang serologist forensik menganalisa berbagai
cairan tubuh termasuk salivam semen, urin dan darah

Karakteristik Darah

Plasma is the fluid portion of the


blood (55%)
Cells (45%)
Erythrocytes are red blood
cells. They are responsible for
oxygen distribution.
Leukocytes are the white blood
cells; they are responsible for
cleaning the system of
foreign invaders.
Thrombocytes or platelets are
responsible for blood clotting
Serum is the liquid that separates
from the blood when a clot is
formed.
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Historical Perspective
of Blood Typing

Around 1900, Karl Landsteiner discovered that there are


four different types of human blood based on the
presence or absence of specific antigens found on the
surface of the red blood cells.
In 1940, Landsteiner and Weiner reported the discovery
of the Rh factor by studying the blood of the Rhesus
monkey. 85% of Caucasians, 94% of Black Americans
and 99% of all Asians are Rh positive.

Blood Terminology
ABO blood groupsbased on having an A, B, both or no
antigens on red blood cells
Rh factormay be present on red blood cells; positive if
present and negative if not
Antigen chemical characteristics (proteins) on the surface
of red blood cells. These account for blood type.
Antibodya substance that reacts with an antigen. Found in
the serum.
Agglutinationclumping of red blood cells; will result if blood
types with different antigens are mixed

Presumptive Tests for


Blood Determination
Kastle-Meyer color testa mixture of
phenolphthalein and hydrogen peroxide; the
hemoglobin will cause the formation of a deep pink
color if blood is present
Hematest stripsreacts with the heme group in
blood causing a blue-green color
Luminol testreaction with blood to produce light
Capable of detecting bloodstains diluted up to 300,00
times. Reveals blood patterns that may have gone
unoticed.
Does not interfere with DNA testing.
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Luminol Test

Human vs Animal Blood

Microscopic observation
Precipitin testblood is injected into a rabbit;
antibodies are formed; the rabbits blood is
extracted as an antiserum; the antiserum is
placed on sample blood. The sample will react
with human proteins if human blood is present.
This test is very sensitive and requires only a
small amount of blood.

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Precipitin Test

Animal Blood

Larger nucleic red


blood cells

Frog Blood

Human Blood
Red blood cells are most
numerous; 5 to 6 million
per mm3
White blood cells are
larger and less numerous;
5 to 10,000 per mm3
Platelets are tiny, cellular
fragments; 350 to 500,00
per mm3

13

Blood Typing
Blood type A has antigen A on the surface of
the cell and will agglutinate with antibody A.
Blood type B has antigen B on the surface of
the cell and will agglutinate with antibody B.
Blood type AB has antigens A and B on the
surface of the cells and will agglutinate with
both antibody A and B.
Blood type O has neither antigen A or B and
will not agglutinate.
Antibodies have two reactive sites (bivalent)

Blood Groups
Antibody Can Give

Can Get
Blood From

Type

Antigen

A,AB

O,A

B,AB

O,B

AB

AandB

Neither
AnorB

AB

A,B,O,AB

Neither
AnorB

AandB

A,B,O,AB

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Blood To

Blood Typing
Human serum containing specific antibodies
can be purchased
Separately mix a drop of unknown blood
sample with drop of each anti-serum

Reaction between blood


and anti-serum
No reaction between
blood and anti-serum

Blood Typing Example

A sample of unknown blood is mixed with three anti-sera samples:

Tube 1 (Anti-A): No reaction


Tube 2 (Anti B): No reaction
Tube 3 (Anti Rh): Cloudy reaction
In terms of the A-B-O and Rh systems, what type blood is the
sample?
O positive

Secretors

80% of the population are secretors.


Their blood-type antigens are found in
high concentration in their body fluids
such as saliva, semen, vaginal
secretions and gastric juice.

18

Genetics of blood
Genes are composed of alleles. Blood
type has 3 alleles: A,B,O.
The allele is responsible for producing its
corresponding antigen. The O allele
produces no antigen.

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Genetics of Blood
Genotype - the possible allele combinations
from the parents
Phenotype - the blood type the child actually
has
The A and B alleles are dominant over the O
allele as the O allele doesnt produce an
antigen.
The A and B alleles are codominant as both
antigens are produced.
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Punnet Squares
Assume a set of parents with a father
phenotype A and genotype AO; and a
mother of phenotype B and genotype BO.
The child must get one allele from each
parent. What are the possible genotypes
and phenotypes.

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Mother

Father
A
O
B AB BO
O AO OO

Genotypes
Phenotypes - AB, B, A, O

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Blood Types and the


Population
O positive is the most common blood type. Not all ethnic groups have the same mix of
these blood types.
Hispanic people, for example, have a relatively high number of Os, while Asian people

have a relatively high number of Bs

Rhesus Rh Blood Types


Rh blood types were discovered in 1940 by Karl Landsteiner and
Alexander Wiener. This was 40 years after Landsteiner had discovered
the ABO blood groups.
The Rh system was named after rhesus monkeys, since they were
initially used in the research to make the antiserum for typing blood
samples. If the antiserum agglutinates your red cells, you are Rh+ . If
it doesn't, you are Rh- . Despite its actual genetic complexity, the
inheritance of this trait usually can be predicted by a simple conceptual
model in which there are two alleles, D and d. Individuals who are
homozygous dominant (DD) or heterozygous (Dd) are Rh+. Those who
are homozygous recessive (dd) are Rh- (i.e., they do not have the key Rh
antigens).

Clinically, the Rh factor, like ABO factors, can lead to serious medical
complications. The greatest problem with the Rh group is not so much
incompatibilities following transfusions (though they can occur) as those
between a mother and her developing fetus.
Mother-fetus incompatibility occurs when the mother is Rh- (dd) and
the father is Rh+ (DD or Dd). Maternal antibodies can cross the
placenta and destroy fetal red blood cells. The risk increases with each
pregnancy. Europeans are the most likely to have this problem--13% of
their newborn babies are at risk.
Rh type mother-fetus incompatibility occurs only when an Rh+ man
fathers a child with an Rh- mother. Since an Rh+ father can have either a
DD or Dd genotype, there are 2 mating combinations possible:

Rhesus Rh Blood Types

Only the Rh+ children (Dd) are likely to have


medical complications. When both the mother and
her fetus are Rh- (dd), the birth will be normal.

Human fetus in a mother's


uterus (the umbilical cord and
placenta connect the fetus to its
mother)

Rh antibodies are harmless until the mother's second or later pregnancies. If she is ever carrying another Rhpositive child, her Rh antibodies will recognize the Rh proteins on the surface of the baby's blood cells as
foreign, and pass into the baby's bloodstream and attack those cells. This can lead to swelling and rupture of
the baby's RBCs. A baby's blood count can get dangerously low when this condition, known as hemolytic or
Rh disease of the newborn, occurs

Rhesus Rh Blood Types

If a father's Rh factor
genes are + +, and the
mother's are + +, the
baby will have one +
from the father and
one + gene from the
mother. The baby will
be + + Rh positive.

If a father's Rh factor
genes are + +, and
the mother's are - -,
the baby will have
one + from the father
and one - gene from
the mother. The baby
will be + - Rh
positive.

If the father's
genes are - -, and
the mother's are -, the baby will
be: - - Rh
negative

If the father's genes


are + - Rh positive,
and the mother's are +
- Rh positive, the
baby can be:
+ + Rh positive
+ - Rh positive
- - Rh negative

If the father's genes are -, and the mother's are +


-, the baby can be:
+ - Rh positive
- - Rh negative

Why is Rh disease a concern?


When an Rh negative mother has a baby that is Rh positive, problems can
develop if the baby's red blood cells cross to the Rh negative mother. This
usually happens at delivery when the placenta detaches. It may also
happen, however, anytime blood cells of the two circulations mix such as
during a miscarriage or abortion, with a fall, or during an invasive prenatal
testing procedure such as an amniocentesis or chorionic villus sampling.
The mother's immune system sees the baby's Rh positive red blood cells as
foreign. Just as when bacteria invade the body, the immune system responds
by developing antibodies to fight and destroy these foreign cells. The
mother's immune system keeps the antibodies in case the foreign cells
appear again, even in a future pregnancy. The mother is now Rh sensitized.
Although it is not as common, a similar problem of incompatibility may
happen between the blood types (A, B, O, AB) of the mother and baby in the
following situations:

Mother's Blood Type O A B Baby's Blood Type A or B B A In a first pregnancy,


Rh sensitization is not likely. Usually it only becomes a problem in a future
pregnancy with another Rh positive baby. During that pregnancy, the mother's
antibodies cross the placenta to fight the Rh positive cells in the baby's body. As
the antibodies destroy the red blood cells, the baby can become anemic. The
anemia can lead to other complications including jaundice and organ
enlargement.
Rh disease is also called erythroblastosis fetalis during pregnancy. In the
newborn, the resulting condition is called hemolytic disease of the newborn
(HDN).
Some of the more common complications of Rh disease for the fetus and newborn
baby include the following:

anemia (in some cases, the anemia is severe with enlargement of the liver and spleen)
jaundice - yellowing of the skin, eyes, and mucous membranes.
severe anemia with enlargement of the liver and spleen
hydrops fetalis - this occurs as the fetal organs are unable to handle the anemia. The heart begins to
fail and large amounts of
fluid build up in the fetal tissues and organs. A fetus with hydrops
fetalis is at great risk of being stillborn.

After birth, the red blood cell destruction may continue.


Problems may include the following:
severe jaundice
The baby's liver is unable to handle the large amount of a substance
called bilirubin that results from red blood cell. breakdown. The
baby's liver is enlarged and anemia continues.
kernicterus
The most severe form of too much bilirubin and results from the build
up of bilirubin in the brain. This can cause seizures, brain damage,
deafness, and death

Population data
The frequency of Rh factor blood types and the RhD neg allele gene differs in various
populations.
Population data for the Rh D factor and the RhD neg allele[

BLOOD PATTERN/SEROLOGICAL
RECONSTRUCTION

SCENE PATTERN
RECONSTRUCTION

LAB RESULTS
RECONSTRUCTION

1.
2.
3.
4.
5.

1.
2.
3.
4.
5.

Stain condition
Pattern
Distribution
Location
Directionality

Genetic marker typing


Age Determination
Source Determination
Race Determination
Sex Determination

FromCrackingCasesbyDr.HenryC.Lee

32

BLOOD DROPLET
Characteristics
A blood droplet will remain spherical in
space until it drops onto a surface
Once a blood droplet impacts a surface,
a bloodstain is formed.
A droplet falling from the same height,
hitting the same surface at the same
angle, will produce a stain with the
same basic shape.
33

CONDITIONS AFFECTING
BLOODSTAIN SHAPE
Size of the droplet
Angle of impact
Velocity at which the blood droplet left the
original surface
Texture of the target surface
On clean glass or plastic--droplet will have smooth outside
edges
On a rough surface--will produce scalloping on the edges

34

Questions Answered by
Blood Spatter Interpretation
The distance between the target
surface and the origin of blood at the
time of blood shed
The point(s) of origin of the blood
Movement and direction of a person or
an object
The number of blows, shots, etc.
causing the bloodshed and/or the
dispersal of blood.
35

Questions Answered by
Blood Spatter Interpretation
Type and direction of impact that
produced the bloodshed
The position of the victim and/or object
during bloodshed
Movement of the victim and/or object
after bloodshed

36

Bloodstain Terminology
Angle of impact--angle at which blood strikes a
target surface.

Bloodstain transfer--When a bloody object comes


into contact with a surface and leaves a patterned blood
image on the surface.

Backspatter--blood that is directed back toward its


source of energy.

Cast-off--blood that is thrown from an object in


motion

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Bloodstain Terminology
Contact stain--general term referring to bloodstains

caused by contact between a wet, blood-bearing surface


and a second surface which may or may not have blood
on it
Transfer--image is recognizable and may be
identifiable with a particular object
Swipe--wet blood is transferred to a surface which
did not first have blood on it
Wipe--a non-blood bearing object moves through a
wet bloodstain, altering the appearance of the
original stain
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Bloodstain Terminology
Directionality--relates to the direction a drop of
blood traveled in space from its point of origin

Terminal velocity--the greatest speed to which


a free falling drop of blood can accelerate in air. It is
dependent upon the acceleration of gravity and the
friction of the air against the blood--approximately
25:1 feet/second.

High velocity--greater than 100 feet per second;


gives a fine mist appearance

Low velocity--5 feet per second or less


Medium velocity--5 to 25 feet per second.
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Bloodstain Pattern

Terminal Velocity
Directionality
Angle of Impact

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Blood Stain Patterns


The shape of a blood stain:

Round--if it falls straight down at a 90


degree angle.
Elliptical--Blood droplet elongates as the
angle decreases from 90 to 0 degrees.
The angle can be determined by the
following formula:
width
=
sine of the impact angle
length
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IMPACT
The more acute the angle of impact, the
more elongated the stain.
90 degree angles are perfectly round
with 80 degree angles taking on a more
elliptical shape.
At about 30 degrees the stain will begin
to produce a tail.
The more acute the angle, the easier it
is to determine the direction of travel.
42

Blood Splatter Analysis


Categories of Bloodstains:

Passive (dripping)
Transfer (smearing)
Projected

Projected Bloodstains occur in shootings, trauma from blunt


weapons, hacking, or slashing attacks.

Blood Stains
The harder and less porous
the surface, the less the blood
drop will break apart.
The softer and more porous
the surface, the more a blood
drop will break apart.
The pointed end of the blood
stain faces the direction the
stain is traveling.
44

Projected Bloodstain Analysis


Two Important Determinations:
a. direction of splatter

b. angle of impact with surface


sin = (width drop / length drop)

Area of Intersection
The location of the blood
source can be
determined by drawing
lines the various blood
droplets to the point
where they intersect.
This is the bloods
origin.
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Area of Convergence
The area of
convergence is the
point of origin; the
spot where the
blow occurred.
It is determined by
drawing a line from the
area of intersection
straight up to where to
where the angle of
impact would intersect

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Uses of Blood
Spatter
There are many uses for blood spatter. It can be used to find the type of weapon
and the velocity the victim was struck with it. It can also tell us the number of
blows given, the handedness of the assailant, and the position and movements
of both the victim and the assailant during and after the attack. As well as the
type of injuries, how long ago the crime took place, and whether the death of
the victim was immediate or delayed.

Advantages - Blood spatter can be used to a murder teams advantage. It can


help determine variables that may not be evident without an autopsy.
Detectives will have some information much faster, such as how long ago the
crime was committed and whether the death was immediate or not. It can
provide evidence that may disprove any lies a suspect may tell. Which wounds
were inflicted when, for example, or the position the assailant found himself in.
It can also help narrow down suspects by revealing the handedness of the
assailant. The absence of blood can also help to solve a crime. Void patterns
are areas where blood should be, but isnt. Often it can tell detectives where
the assailant was standing.

Disadvantages - Of course there are also


disadvantages to blood spatter. If there
are multiple victims or assailants it may
get complicated and it can be difficult to
determine what exactly happened. There
is also the fact that not every crime
involves blood or blood spatter at all. This
renders blood spatter analysts useless for
the time being.

CRIME SCENE

Whatevidencecan
youseeinthis
crimescene?What
storydoesthescene
tell?

55

Bring In The Dogs!


Thedoglocates
humanscent.A
closerlookshows
thattheasheshave
humanremainsand
clothing.In
addition,lookclosely
attherocksonthe
nextslide.

56

Blood Evidence
Class evidence for blood would include blood
type. If you can determine the DNA you
would have individual evidence.
Blood stain patterns are considered
circumstantial evidence in a court room.
Experts could argue many points including
direction of stains, height of the perpetrator,
position of the victim, left/right hand, whether
the body was moved, etc.

58

Forensic Characterization of
Semen
Many crimes involve sexual misconduct
Normal males, upon ejaculation, release seminal fluid. This fluid
is a mixture of components:
water
spermatozoa
enzymes
inorganic salts
Semen at a crime scene (clothing, bed sheets, carpet, cushions)
can be located and identified by three common methods:
microscope examination to find sperm
acid phosphatase test
p30 (also called prostrate specific antigen) immunoassay test

Sperm
Among the smallest and most
highly specialized cells in the
human body.
Has a head and a tail
Contains 23 chromosomes
with the genetic material
found in the head
Males release 2.5 to 6
milliliters of seminal fluid per
ejaculation with approximately
100 million sperm per milliliter.

Magnified400X
60

Semen
Determination of Seminal Fluid
Acid phosphatase color test
the presence of acid phosphatase, the
enzyme secreted by the prostate gland into
the seminal fluid, will turn purple when
sodium alpha naphthylphosphate and Fast
Blue B solution are placed on it.
It will also fluoresce under UV light when it
comes in contact with 4-methyl umbelliferyl
phosphate.
61

Semen (cont.)
Determination of Seminal Fluid
Prostate Specific Antigen (PSA) or p30-unique to seminal plasma
P30 is isolated and injected into a rabbit where
antibodies are produced (anti-p30)
The stain extract is place in one well of an
electrophoresis plate and the anti-p30 in the
opposite well. The electric is applied and the
antigens and antibodies move toward each other.
The formation of a precipitation line between the
wells shows the presence of p30 in the sample
stain. It must be seminal fluid.
62

Testing for Seminal


Stains
Semen can be unequivocally identified by either the presence of
spermatozoa or of p30, a protein unique to seminal plasma.

Forensic scientists can successfully link seminal material to an individual by


DNA typing.

Forensic Characterization of
Saliva
Saliva is a mixture of many components:
99% water
Mucin (protein helps in swallowing)
Amylase (enzyme to help digest carbohydrates)
Cheek cells (good for DNA)

Adults produce 1.0-1.5 liters of saliva/day and it is not


uncommon at crime scenes (especially involving bite marks).
A simple test for saliva involves mixing
starch, iodine, and a sample of the
presumed saliva together. Starch and
iodine are a deep blue color when mixed
together. The amylase breaks down
starch, however, and the color fades
(takes about 15 mins @ 37 oC).

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