1
Forensic Serology
WHAT IS IT?
Serology - is the scientific study of
blood serum.

Purposes: Medical or Forensic

Blood samples can change the entire case

1930, Karl Landsteiner received a Noble Prize award for
his research in serology. He introduced to the scientific
world that all human blood can be grouped into 4 types
known as O, A, B, AB.
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Forensic Serology
Serological tests may be performed for diagnostic purposes
when an infection is suspected, in rheumatic illnesses, and
in many other situations, such as checking an individual's
blood type
Diagnostic ID of antibodies in the serum.
 Antibodies are formed in response to an infection
(against a given microorganism)
 Against other foreign proteins (in response, for example,
to a mismatched blood transfusion)
 One's own proteins (in instances of autoimmune
disease).
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Forensic Serology

Serology techniques vary depending on the antibodies
being studied. Some are: ELISA, agglutination, precipitation,
complement-fixation, and fluorescent antibodies.

Serological tests are not limited to blood serum, but can also
be performed on other bodily fluids such as semen & saliva,
which have similar properties to serum.

Serological tests may also be used forensically, generally to
link a perpetrator to a piece of evidence (e.g., linking a rapist
to a semen sample).
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Forensic Serology
The Crime Scene
and Blood

Wet Blood vs. Dried Blood

Blood found in different forms

Refrigerated red blood cells and serum from white blood cells

Where DNA can be taken from

Blood pattern analysis helps in the reconstruction of many
crime scenes
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Forensics of Blood
The criminalist needs to answer the following
questions when examining dried blood:


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Is it blood?
From what species did the blood originate?
If the blood is of human origin, how closely can
it be associated to a particular individual?
The determination of blood is best made by
6 means of a preliminary color test.
Blood (HEMO)



Blood: water, white blood
cells(leukocytes), red blood cells
(erythrocytes), various
platelets(last component of blood),
various proteins, plasma.
Red blood cell : RBCs, Carry
oxygen to organs of the body, O2.
White blood cells: WBCs, part of
the Immune system, protects your
body from infection and diseases
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Blood (HEMO)

The cellular fraction of blood contains red blood
cells (erythrocytes) and white blood cells
(leucocytes)
RBC

White blood cells (WBC) are
the source of DNA for DNA
typing analysis
WBC

Red blood cells (RBC) do not
contain any nuclear DNA
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Plasma cells

Plasma: primary component (55%) in blood,
it has two component:


Serum- yellow liquid, contains specific antibody
Clotting Proteins
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Blood and Body Fluid Individuality:
Traditional Approaches
1. The Classical or Conventional Genetic Markers:

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blood groups
isoenzymes
plasma (serum) proteins
hemoglobin variants
HLA
The first blood group markers were ABO, discovered
in 1901 by Karl Landsteiner
Blood and Body Fluid Individuality:
Traditional Approaches

ABO markers were first applied to criminal cases
involving bloodstains by Dr. Leon Lattes of Italy in
1913

Isoenzymes are enzymes which occur in multiple
molecular forms, reflecting differences in the gene
that code for the enzyme

Similarly, there are common variants of the protein
hemoglobin
Blood and Body Fluid Individuality:
Traditional Approaches
2. How Does Typing Genetic Markers Help
“Individualize” a Biological Specimen?
 A gene is a region of DNA that codes for a
particular protein or enzyme

Because chromosomes are paired (maternal and
paternal), and there is one gene on each
chromosome, the genes are paired

A gene locus is the location on a chromosome
where a particular trait is determined
Blood and Body Fluid Individuality:
Traditional Approaches

The genes making up a pair at a given locus are
called alleles

The alleles may be the same (homozygous) or
different (heterozygous)

Population genetics looks at how often alleles found
at a given locus occur in a population
Blood and Body Fluid Individuality:
Traditional Approaches

A portion of a large population is sampled and
tested to determine the frequency of a particular
allele

Statistics are used to estimate the frequency of an
allele in the entire population
Antibodies and Antigens
Antibody: Proteins that destroy or inactivate a
specific antigen and are found in serum.
Antigen: Substances that stimulates the
proteins to produce
antibodies against it.
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A-B-O vs DNA

Prior to the advent of DNA typing, bloodstains were
linked to a source by A-B-O typing and the
characterization of polymorphic blood enzymes
and proteins.

This approach has now been supplanted by the
newer DNA technology.
DNA analysis has allowed forensic scientists to
associate blood and semen stains to a single
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individual.

Forensic Serology
Preserving Evidence at the
Crime Scene

Blood evidence results if prepared and collected right

Communication from beginning of crime scene to court

Videotaping Blood Evidence

What Type of Film to use and at medium range

Chain of Custody
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Forensic Serology
 A term which describes laboratory tests
which employ a specific antigen/antibody
reactions. The science dealing with serums
and especially their reactions and
properties.
 It includes blood splatter patterns that leads
to crime-scene analysis and eventually to a
criminal.
 Every species has a unique type of blood.
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Forensic Identification of Blood
Categories of identification tests:

Presumptive or preliminary test



Used for screening specimens that might contain the
substance or material of interest
Both false positive and false negative results may be
obtained
Confirmatory test


Are tests which are entirely specific for the substance or
material for which it is intended
A positive confirmatory test is interpreted as an
unequivocal demonstration that the specimen contains the
substance or material
Forensic Identification of Blood
Presumptive Tests for Blood:
 Presumptive blood tests are used to screen evidence for the
possible presence of blood
 Most are color tests and are based on the peroxidase-like
activity of hemoglobin
 Peroxidase catalyzes the following reaction
 Reduced Dye + peroxide --> Oxidized dye + water
 The presence of hemoglobin catalyzes the reaction, forming
a colored dye product
 Positive presumptive tests do not prove that blood is present
Forensic Identification of Blood
Confirmatory Tests for Blood:
 Older tests included crystal tests such as the
Teichmann and Takayama tests
 Current immunological tests use antibodies
specific for human hemoglobin, thus combining
the confirmatory test for blood with a human
species test
 The crystal tests and the immunological tests are
known as direct confirmatory tests
Species Determination
Tests must be done on blood specimens to determine
the species of origin
 Species origin tests are done using immunological
methods which involve the interaction of antigens and
antibodies
 Hemoglobin from human red blood cells can be used
as the antigen to produce anti-human hemoglobin
serum
 Specific antiserum can be used to test for the
presence of antigens in unknown specimens
Species Determination



Common immunological species tests include the
Ouchterlony method
Extracts of the bloodstain to be analyzed are tested with
specific antisera
If the bloodstain contains the antigens corresponding to the
specificity of the antiserum, a visible precipitate (precipitin)
is obtained
Blood is the most common and
most important evidence in the
world of criminal justice.
Forensic Nature of Blood
Stains
http://projects.nfstc.org/gallery/ma
in.php?g2_itemId=3857
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General Bloodstain Features

Bloodstain patterns deposited on floors, walls,
bedding, and other relevant objects can provide
valuable insights into events that occurred during
the commission of a violent crime.

The crime scene investigator must remember that
the location, distribution, and appearance of
bloodstains and spatters may be useful for
interpreting and reconstructing the events that
produced the bleeding.
General Bloodstain Features
The information one can uncover as a result of
bloodstain pattern interpretation includes:


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The direction from which blood originated
The angle at which a blood droplet struck a surface
The location or position of a victim at the time a bloody
wound was inflicted
The movement of a bleeding individual at the crime
scene
The approximate number of blows that struck a bleeding
victim
The approximate location of an individual delivering
blows that produced a bloodstain pattern
The Tests

A positive result from the KASTLE-MEYER color
test is highly indicative of blood.
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A presumed blood sample is first collected with a
swab.
A drop of phenolphthalin reagent is added to the
sample, and after a few seconds, a drop of
hydrogen peroxide is applied to the swab.
Hemoglobin causes a deep pink color. If the swab
turns pink rapidly, it is said to test presumptive
positive for blood.
Waiting for periods over 30 seconds will result in
most swabs turning pink naturally as they oxidize
on their own in the air.
The Tests

The LUMINOL TEST is used to search out
trace amounts of blood located at crime scenes.

Produces light (luminescence) in a darkened area. To exhibit its
luminescence, the luminol must first be activated with an oxidant
(H2O2) . Usually, a solution of hydrogen peroxide (H2O2) and a
hydroxide (OH-) salt in water is used as the activator. In the
presence of a catalyst such as an iron compound (Heme-in
blood), peroxide is decomposed to form oxygen and water: 2
H2O2 → O2 + 2 H2O
In the forensic detection of blood, the catalyst is the iron present
in hemoglobin. Enzymes in a variety of biological systems may
also catalyze the decomposition of hydrogen peroxide.

BlueStar™ TEST- Comm. Test avail. – doesn’t need total
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darkness to observe in the field
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The Tests

Precipitin Test – simple test to determine if blood is of
human or animal origin. (When animals are injected with
human blood, antibodies form reacting with the human
blood forming a precipitin band.

Microcrystalline tests, such as the Takayama and
Teichmann tests, depend on the addition of specific
chemicals to the blood so that characteristic crystals will be
formed. It is a less sensitive test for blood ID.

Once it has been determined that the bloodstain is of
human origin, an effort must be made to associate or
dissociate the stain with a particular individual.
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Blood Stain Pattern Analysis


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Combination of physics,
hydraulics, and common
sense
Helps to reconstruct
crime scene and identify
criminals
Draws from the sciences
of biology, chemistry,
math, and physics




What mechanism
creates stains
What direction a
droplet of blood was
traveling
Type of object used in
attack
Position of victim, 30
suspect, and objects
Blood Spatter

Blood Spatter is called Bloodstain Pattern
Interpretation.

It involves reconstructing the events that must
have happened to produce the bleeding.

It is not something that most law enforcement
officials can do--it usually requires a specialist.
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Blood Spatter
Herbert L. MacDonell published a study in 1971; gave tips to
investigators:
 To determine the impact angle of blood on a flat surface by
measuring the degree of circular distortion of the stain. The
more the angle decreases, the more the stain is less circular
and more oblong.
 Surface texture is one of the key components in determining
spatter type, it means that the harder the surface is, the less
spatter will result.
 When a droplet of blood hits a surface which is hard as well
as smooth, the blood usually breaks apart upon impact. This
in turn causes smaller droplets. These droplets will continue
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to move in the same direction as the original droplet.
Effects of Surface Texture

Surface texture and the stain’s shape, size,
and location must be considered when
determining the direction, dropping distance,
and angle of impact of a bloodstain.

Surface texture is of paramount importance.
In general, the harder and less porous the
surface, the less spatter results.
Directionality and Angle of Impact

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The direction of travel of blood striking an object may be
discerned because the pointed end of a bloodstain
always faces its direction of travel.
The impact angle of blood on a flat surface can be
determined by measuring the degree of circular
distortion. At right angles the blood drop is circular, as
the angle decreases, the stain becomes elongated.
Mathematically, the angle of impact can be calculated by
the equation:
Sin A = Width of blood stain
Length of blood stain
Impact Bloodstain Spatter
Patterns
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Impact spatter occurs when an object
impacts a source of blood.
Forward spatter is projected outward and
away from the source.
Back spatter, sometimes called blow-back
spatter, is projected backward from the
source,
Classifying Impact Spatter
Using droplet size to classify impact patterns by velocity gives
investigators insight into the general nature of a crime but
cannot illuminate the specific events that produced the
spatter pattern.
 Low Velocity Spatter - drops with diameters of 3 mm or
more normally produced by minimal force at a speed of
less than 5 ft/sec.
 Medium Velocity Spatter - drops with diameters from 1-3
mm at a speed of 5 to 25 ft/sec.
 High Velocity Spatter - drops with diameters of less than
1mm at a speed of 100 ft/sec or faster.
1. Determination of Angle of blood
 The drop on the right fell 42 inches at an
angle of 60-degrees on toward smooth
cardboard, while the drop on the left fell the
same 42 inches and on to the same
surface, but at a 10-degree angle.
Flight Characteristics and Stain Patterns of Human Blood, Herbert L.
MacDonnell (Washington DC: USGPO,1971), pp. 44,49
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Blood Spatter

The blood drop at the right struck a plastic wall tile
falling again from 42 inches. On the left, the drop
fell from the same distance on to a piece of heavy,
unusual textured wallpaper.
Flight Characteristics and Stain Patterns of Human Blood, Herbert L.
MacDonnell (Washington DC: USGPO,1971), pp. 36,37
38
Blood Spatter

This drop of blood was tested traveling 4ft/sec.,
on to a hard cardboard surface. It was traveling
from right to left at an angle of 56-degrees.
Although not clear in the example, MacDonnell's
test showed the smaller drops aimed toward the
large one.
Flight Characteristics and Stain Patterns of Human Blood, Herbert L.
MacDonnell (Washington DC: USGPO, 1971), pg. 39
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Area of Convergence

The Area of Convergence is the point on a
two-dimensional plane from which the drops
in an impact pattern originated.

This can be established by drawing straight
lines through the long axis of several
individual bloodstains, following the line of
their tails.
Area of Origin

The Area of Origin of an impact bloodstain
pattern is the area in a three-dimensional space
from which the blood was projected.

This will show the position of the victim or suspect
in space when the stain-producing event took
place.

The string method is commonly used at a crime
scene to approximate the position of the area of
origin using found angles of impact of individual
stains in the pattern.
Gunshot Spatter

Gunshot spatter is characterized by fine forward
spatter from an exit wound and back spatter from
an entrance wound. However, the gunshot
produces only back spatter if the bullet does not
exit the body.

Depending upon the distance from the victim that
the gun was discharged, some back spatter may
strike the gunman and enter the gun muzzle. This
is called the drawback effect.
Cast-off Spatter

A Cast-off pattern is created when a blood-covered
object flings blood in an arc onto a nearby surface.

This kind of pattern commonly produced by a bloody
fist or weapon between delivering blows.

The features of the cast-off pattern are affected by the
size of the object, the amount of blood, and the
direction the object was moving.

By counting and pairing forward/backward patterns,
one may determine the minimum number of blows
delivered.
Arterial Spray Spatter

Arterial spray spatter is created when a victim
suffers an injury to a main artery or the heart, and
the pressure of the continuing pumping of blood
causes blood to spurt out of the injured area.

The site of the initial injury to the artery can be
found where the pattern begins with the biggest
spurt. The trail away from this point shows the
victim’s movement.

The oxygenated blood spurting from the artery
tends to be a brighter red color than blood
expelled from impact wounds.
Expirated Blood Patterns

An Expirated Blood Pattern is created by blood that
is expelled from the mouth or nose from an internal
injury.

The presence of bubbles of oxygen in the drying
drops or a lighter color as a result of dilution by saliva
can differentiate a pattern created by expirated blood.

The presence of expirated blood gives an important
clue as to the injuries suffered and the events that
took place at a crime scene.
Void Patterns

A void is created when an object blocks the deposition
of blood spatter onto a target surface or object and the
spatter is deposited onto the object or person instead.

The blank space on the surface or object may give a
clue as to the size and shape of the missing object or
person.

Voids may be applicable for establishing the body
position of the victim or assailant at the time of the
incident.
Contact/Transfer Patterns

A Contact or Transfer Pattern is created when
an object with blood on it touches one that does
not have blood on it.

Simple transfer patterns are produced when the
object makes contact with the surface and is
removed without any movement of the object.
Contact/Transfer Patterns

The size and general shape of the object may be
seen in a simple transfer.

Other transfers may be caused by movement of
the bloody object across a surface.
Flows

Flow patterns are made by drops or large
amounts of blood flowing by the pull of gravity.

“Active” flows are formed by blood coming from an
actively bleeding wound.

“Passive” flows originate with blood deposited on a
surface such as an arterial spurt.
Flows

The flow direction may show movements of objects
or bodies while the flow was still in progress or
after the blood dried.

Interruption of a flow pattern may indicate the
sequence and passage of time between the flow
and its interruption.
Pools

A pool of blood occurs when blood collects in a
level and undisturbed place.

Blood that pools on an absorbent surface may be
absorbed throughout the surface and diffuse.

The approximate drying time of a pool of blood is
related to the environmental condition of the
scene. By experimentation an analyst may be
able to reasonably estimate the drying times of
stains of different sizes.
Skeletonization

The phenomenon of skeletonization occurs when
the edges of a stain dry to the surface.

This usually occurs within 50 seconds of
deposition of droplets, and longer for larger
volumes of blood.
Skeletonization

After this time, if the bloodstain is altered by a
contact of wiping motion, the skeletonized
perimeter will be left intact.

Knowing the skeletonization time, an investigator
determine the timing of movement or activity.
Trail Patterns

A trail pattern is a series of drops that are
separate from other patterns, formed by blood
dripping off an object or injury.

The stains form a kind of line or path usually
made by the suspect after injuring or killing the
victim or injuring themselves.
Trail Patterns

The pattern may show movement, lead to a
discarded weapon, or provide identification of
the suspect by his or her own blood.

The shape of the stains in trail patter can help
investigators determine the direction and speed
at which a person was moving.
Documenting Bloodstain
Evidence

Investigators should note, study, and
photograph each pattern and drop to accurately
record the location of specific patterns and to
distinguish the stains from which laboratory
samples were taken.

In photographs specific techniques are used to
document bloodstain patterns.


The grid method involves setting up a grid of
squares of known dimensions over the entire pattern.
The perimeter ruler method involves setting up a
rectangular border of rulers around the pattern and
then placing a small ruler next to each stain.
Interpreting Bloodstain
Evidence


Some jurisdictions have a specialist on staff
who will decipher patterns either at the
scene or from photographs at the lab.
However, it is important that all personnel be
familiar with patterns to properly record and
document them for use in reconstruction.
Bloodstain Characterization

Determination of the group and type of
blood, blood testing, bloodstain examination
are the main functions of the forensic
serologist.

In many crime labs, there may be no clear
distinction between job title and job function.
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Rape
Evidence of a sexual assault can come in a
variety of forms
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Rape Evidence
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The rape victim must undergo a medical
examination as soon as possible after the assault.
At that time the appropriate items of physical
evidence including clothing, hairs, and vaginal and
rectal swabs can be collected for subsequent
laboratory examination.
All outer and undergarments should be carefully
removed and packaged separately in paper (not
plastic) bags.
Bedding, or the object upon which the assault took
place, may also be carefully collected.
Rape Evidence

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If a suspect is apprehended within 24 hours
of the assault, it may be possible to detect the
victim’s DNA on the male’s underwear or on a
penile swab of the suspect.
Items routinely collected from the suspect
include all clothing, pubic hair, head hair,
penile swab, and a blood sample or buccal
swab for DNA typing.
The forceful physical contact between victim
and assailant may result in a transfer of such
physical evidence of blood, semen, saliva,
hairs, and fibers.
Collection of Rape Evidence

Lack of presence of semen does
not rule out a sexual assault.

Physical injuries such as
bruises, bleeding, or strangle
marks can be indicators.

As Locard stated with his theory,
physical contact may result in a
transfer of physical evidence
such as hair, fibers, blood, and
semen.

DNA can be collected
from everywhere on
the human body
including dandruff and
fingernails.

To protect this kind of
evidence it is important
to properly remove,
package, and store this
evidence.
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Collection of Rape Evidence

It should be stored
separately in paper bags.

Even the victim can be
wrapped in something like a
cloth if they haven’t washed
off any potential evidence.

The bed or surface on
which the assault took place
can also be collected as
evidence.

The collector of the
evidence should be
wearing latex gloves to
avoid contamination of
evidence.

The rape victim must
have a medical
examination done as
soon as possible with a
sexual assault evidence
kit.
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Collection of Rape Evidence

A thorough examination will
consist of collecting the
following: Pubic combings,
pubic hair reference samples,
external genital dry skin
areas, vaginal swab, cervix
swab, rectal swab (if the
situation calls for it), oral
swab, head hairs, blood
sample, fingernail scrapings,
all clothing that was worn,
and a urine specimen.
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Collection of Rape Evidence

If a suspect is found, they will be tested for
the following: all clothing that is believed to
have been worn at the time of the assault,
pubic hair combings, pulled head and pubic
hair reference samples, penile swab within
24 hours of assault, and a blood sample.
PUBIC HAIR
PULLED HEAD
HAIR
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Forensic Characterization of
Semen
•The average male releases 2.5 to 6ML of
seminal fluid during an ejaculation. Each
milliliter contains 100 million or more
sperm.
•Examinations of articles that contains
seminal stains can be broken up into two
processes: locating the stain, and proving
it’s identity.
•Often seminal stains are very visible
because they exhibit a stiff, crusty
appearance. However, one can’t always
rely on appearance alone since the stain
isn’t always in an obvious area.
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Forensic Characterization of
Semen
If the fabric has been
washed or contains
minimal amounts of
semen, visual
examination of the
article offers little chance
of detecting the stain.
The best way to locate and
characterize a seminal
stain is to perform the
acid phosphatase color
test.
•
Acid phosphatase is an
enzyme that is secreted
by the prostate gland
into seminal fluid. Its
presence can be easily
detected when it comes
into contact with a
certain acidic solution
and put under a UV light.
67
Forensic Characterization of
Semen
•
•
Sometimes sperm can be
seen through a microscope,
one would think that with the
amount of sperm in a
milliliter they would be easy
to locate but that’s not
always the case.
They are extremely brittle
when dry and disintegrate
easily if the stain is washed
or when the stain is rubbed
against another object which
could happen easily.
•
In the event that there is
no detectable sperm in a
stain, there’s another
way to tell that the stain
is indeed seminal. They
look for a protein called
p30 which is unique to
seminal plasma.
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Testing for Seminal Stains

Many of the cases sent to a forensic laboratory
involve sexual offenses, making it necessary to
examine exhibits for the presence of seminal
stains.

The best way to locate and at the same time
characterize a seminal stain is to perform the acid
phosphatase (an enzyme secreted into seminal
fluid) color test.

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A purple color indicates acid phosphatase enzyme.
Testing for Seminal Stains
70

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 Detection of Semen
Methods of Testing for Semen/Sperm
Acid Phosphatase
Acid phosphatase is an enzyme secreted by the prostate
gland that is present in large amounts in seminal fluid. It can
be found in other biological fluids including vaginal
secretions in smaller quantities.
The PAP assay is a well-documented presumptive assay
for the presence of semen.
Acid phosphatase activity is 50-1000 times greater in
human semen than in any other bodily fluid. Since seminal
and vaginal acid phosphatase can not be discriminated, the
only approach to differentiating semen in vaginal secretion is
by quantitative analysis such as PSA or the microscopic
inspection for sperm.
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Forensic Detection of Semen
Methods of Testing for Semen/Sperm
The advantages of a PSA determination are: ·
Prostate-specific antigen (PSA) or p30 is an accepted
marker for detecting semen in criminal cases.
The PSA test is NOT presumptive like the Acid
Phosphatase (AP) Semen Detection Test.
The detection of PSA is possible in cases where no
spermatozoa can be found (for example vasectomized or
azoospermic men.
PSA can be recovered at detectable concentrations in 30
year old semen stains.
Semen samples can show positive PSA results even at a
dilution factor of 1:200,000.
PSA is detectable in post-ejaculate urine and male urine
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from adult men and can be detected in urine of 11 yr olds.
Forensic Detection of Semen
Methods of Testing for Semen/Sperm
Microscopic Examination for Sperm
There are numerous extraneous objects which are like
the detached heads and tails of spermatozoa as to mislead
even those who are thoroughly experienced in the work.
The generally accepted rule is that no body which simply
resembles the head or tail should be considered as serious
proof and the search must continue until perfectly formed
and entire spermatozoa are recognized.
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Forensic Detection of Semen
Methods of Testing for Semen/Sperm
Microscopic Examination for Sperm

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Forensic Characterization of
Saliva
Consists of more than 99% water
 pH range = 6.8–7.0
 Contains salivary amylase (a digestive enzyme)
 Produced in three main pairs of salivary glands:
parotid, submaxillary, sublingual
Cleanses mouth and provides lubrication
Saliva is always present at the crime scene if
there are bit marks on the victim.
 It can be used to identify an individual through
DNA profiling.
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Blood and Physiological Fluid Evidence:
Evaluation and Initial Examination










How Biological Evidence Analysis Has Changed Because of DNA
Typing
Nature of Blood
Collection, Preservation, and Packaging of Biological Evidence
Test Controls, Substratum Comparison Specimens, and
Contamination Issues
Initial Examination of and for Biological Evidence
Forensic Identification of Blood
Species Determination
Forensic Identification of Body Fluids
Forensic Investigation of Sexual Assault Cases
Blood and Body Fluid Individuality: Traditional Approaches
How Biological Evidence Analysis has
Changed Because of DNA Typing


Prior to the introduction of forensic DNA typing
analysis, blood groups were the genetic markers
that were analyzed from biological evidence
(forensic serology)
Forensic biology now refers to the preliminary
examination of biological evidence prior to the
DNA typing analysis procedures
Collection, Preservation, and
Packaging of Biological Evidence
Blood or Buccal Swabs from Known Person:
 Blood is drawn into a vacutainer tube containing
an anticoagulant such as EDTA (“purple top”
tube)

Buccal (cheek) swabs are
often used in place of liquid
blood as the known sample
Collection, Preservation, and
Packaging of Biological Evidence



Biological Evidence from Scenes:
Fresh or web blood should be collected on clean, sterile,
gauze and allowed to dry
Four sampling methods for dried blood:




Cutting – For stains on objects that are difficult to submit to the lab.
The cut portion should include unstained areas around the
bloodstain
Swabbing – Stain is transferred to a swab which has been
moistened with sterile water or saline.
Scraping – a sharp instrument is used to scrape the stain off of a
surface & onto clean paper
Elution – using a small amount of saline or distilled water to
dissolve the dried stain
Collection, Preservation, and
Packaging of Biological Evidence


The most important consideration for
preserving biological evidence from scenes
is to thoroughly dry the item before
packaging and then store in a cool dry
environment
Biological evidence must be packaged in
paper containers that can breathe
Test Controls, Substratum Comparison
Specimens, and Contamination Issues
1. Known (Exemplar or Reference) Control:


are specimens from a known source
essential for comparison with DNA profiles from
evidentiary specimens
2. Alibi (Alternative) Known Control:

From a known source that may be the source of the
evidence
3. Blank Control:

A specimen known to be free of the item or substance
being tested
Test Controls, Substratum Comparison
Specimens, and Contamination Issues
4. Substratum Comparison Specimens:
 Substratum refers to the underlying material or
surface on which the evidence is found
 A substratum comparison specimen is subjected to
the same testing as the evidence
 The specimen helps to detect interference in lab tests
originating from the evidence surface
 An unstained portion of the evidence underlying
material is collected for this purpose
Test Controls, Substratum Comparison
Specimens, and Contamination Issues
Evidence may be contaminated in several ways:



Biological material may have been on a surface before
the biological evidence was deposited
During scene searching &/or processing activities
During laboratory examinations &/or manipulations
Initial Examination of and for
Biological Evidence
The initial examination is designed to evaluate stains
for possible evidentiary value
Activities include:




Searching for biological stains
Preliminary tests for physiological fluids
Positive preliminary tests are then subjected to
confirmatory tests
Cutting out or transferring stains to swabs for
subsequent examinations
Forensic Identification of Body Fluids
1. Identification of Semen:

Semen is a mixture of specialized cells, called spermatozoa,
suspended in a fluid known as seminal plasma

UV light causes semen stains to fluoresce, and is therefore
used to locate stains

Both presumptive and confirmatory
tests for semen stains are available
Forensic Identification of Body Fluids
Presumptive Test for Semen:
 The AP test is a color test based on the detection of acid
phosphatase, an enzyme from the prostate gland that is
found in high concentration in human semen
Confirmatory Test for Semen:
 A commonly used approach is to use a microscope to detect
spermatozoa in smears made from dried stains
 When no sperm are found, immunological methods are
used to detect the presence of a prostate gland protein
called p30 or PSA
Forensic Identification of Body Fluids
2. Identification of Vaginal Secretions, Saliva, and
Urine:





There are no reliable methods for identifying human vaginal
material
Presumptive tests for saliva are based on the presence of the
enzyme amylase
There are no confirmatory tests for saliva
Presumptive tests for urine are based on the presence of
urea and creatinine
There are no confirmatory tests for urine
Forensic Investigation of Sexual
Assault Cases
1. Coordination of Effort – SANEs and SARTs
 The medical examination of complainants in sexual assault
cases is performed by specially trained sexual assault nurse
examiners (SANE)

Forensic nurses take a lead role in the coordinated
response by the sexual assault response team (SART)

Complainants are taken to a medical facilities or a
SANE/SART facility to attend to their medical needs and to
collect relevant evidence using a sexual assault evidence
collection kit (”rape kit”)
Forensic Investigation of Sexual
Assault Cases
2. The Forensic Scientist’s Role:
 Sexual assault evidence collection kits are forwarded to the
forensic lab for examination

The forensic scientist’s primary role is the analysis of the
physical evidence

If semen is present it helps to establish the corpus delicti
(The material evidence in a homicide, such as the discovered corpse of a murder
victim, showing that a crime has been committed )

If semen or other fluids are found, DNA typing is conducted
to determine if there is a match to a suspect or an exclusion
Forensic Investigation of Sexual
Assault Cases
3. Medical Examination:
 Medical evaluation and treatment of sexual assault
victims initially involves recording the history of the
events, tending to any injuries, and documenting any
injuries, bruises, or contusions

This is followed by evidence collection, which includes
clothing, vaginal swabs, pubic hair combings, any
stains on the skin surface, and a known control (blood
or buccal swab)
Forensic Investigation of Sexual
Assault Cases
4. Sexual Assault Evidence Collection Kits:

Sexual assault evidence collection kits contain a variety of
containers and envelopes plus a detailed set of instructions
on how to use them

Not every container/envelope is used in every case
Forensic Investigation of Sexual
Assault Cases
5. Types of Sexual Assault Cases (3)



unknown offender (identification cases)
known offender (consent cases)
sexual assaults involving children

DNA profiling is helpful in identification cases but
not in consent cases

State laws define the age of consent, thereby
differentiating between an adult and child
Forensic Investigation of Sexual
Assault Cases
6. Drug Facilitated Sexual Assault:
 Commonly encountered “date rape” drugs:



Rohypnol
GHB
Ketamine

Classified as depressants with amnestic effects;
often used along with alcohol

These types of cases require toxicological analysis
of the evidence
General Rules to Consider
- MacDonnell

The smaller the size of bloodspatters, the greater the energy required
to produce them. Low, medium, and high velocity impact spatter may
be identified by their respective sizes but exceptions must also one
considered.

Before a drop of blood can fall, absent any other form of applied
energy, gravitational attraction acting on blood must exceed its surface
tension.

Diameter of a large bloodstain will be of little or not value in estimating
the distance a drop of blood has fallen prior to impact.

When considering the shape of a bloodstain for use in calculating its
angle of impact, only a sharp, well-defined bloodstain should be used
for measuring its width and length.
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General Rules to Consider

Directionality of a blood drop while in flight is usually obvious from the
geometry of its resulting bloodstain. The pointed end indicates the
direction of travel prior to impact on a surface. Directionality may also be
determined when edge scallops appear on only one side of a bloodstain.

Correct interpretation of bloodstain patterns must include consideration of
the surface texture of the material upon which the bloodstains have been
deposited.

Surface tension prevents spattering regardless of the distance a drop of
blood has fallen before impacting a smooth, hard surface such as glass.

Edge characteristics of a bloodstain have no value in establishing the
distance a drop of blood may have fallen prior to impacting a surface
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unless the nature of the surface texture is considered.
General Rules to Consider

Conclusions as to the significance of a limited number of bloodstains
should only be drawn with reservations and/or qualifications. It should be
remembered that a few bloodstains do not a pattern make. There is
nothing wrong with admitting that there are insufficient bloodstains
available to form an opinion. It is far better not to have an opinion than to
have one that is incorrect.

When a dozen or more small bloodstains are present in a recognizable
pattern, their size may allow a prediction as to the energy that was
required to produce them. When the preponderance of individual
bloodstain diameters are less than 1mm, they are consistent with having
been produced as a result of a high velocity impact. Most often they would
result from a shooting.
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General Rules to Consider

When the preponderance of dozens of individual bloodstains are
approximately 1mm or more in diameter, they are consistent with having
been produced as a result of a medium velocity impact. Most often they
would result from a beating or stabbing.

The shape of a bloodstain is a function of the angle at which it impacts a
surface. Perfectly round bloodstains result from a ninety degree impact.
The angle of impact of an elliptical bloodstain may calculated from its
length to width ratio.

Bloodstains may often be lifted from the surface upon which they have
been deposited. The harder and smoother the surface the more likely
they may be successfully lifted.
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General Rules to Consider

When measurements and angles are used to establish the orgin or
origins in space, not only will the actual origin be somewhere below the
point or points of convergence, but it must be remembered that the
investigator is determining a spacial volume and not a small pint of
origin.

Sobriety of the victim will have no significant effect on how bloodstain
patterns are produced. A high blood alcohol level is of no concern to
the interpretation of bloodstain patterns.

When blood is projected upward with sufficient force to strike a ceiling,
it will almost always be the result of a gunshot having an upward
trajectory. Such a trajectory is more often the result of a suicide rather
than it is a homicide.
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