Blood and Bloodstains

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Blood and Bloodstains
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Introduction to Blood
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Introduction to Blood

• What you should already know

Its that red stuff that comes out of your
fingers following a kitchen accident.
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Introduction to Blood
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Blood Facts
The average adult has about FIVE liters of blood
inside of their body, which makes up 7-8 of
their body weight. Blood is living tissue that
carries oxygen and nutrients to all parts of the
body, and carries carbon dioxide and other waste
products back to the lungs, kidneys and liver for
disposal. It also fights against infection and
helps heal wounds, so we can stay healthy. There
are about one billion red blood cells in two to
three drops of blood. For every 600 red blood
cells, there are about 40 platelets and one white
cell.
http//www.bloodbankofalaska.org/about_blood/index
.html
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Introduction to Blood
Red Blood Cells No DNA
White Blood Cells DNA in nucleus
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Introduction to Blood
White Blood Cells DNA in nucleus
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Introduction to Blood

• RBC no DNA WBC yes DNA

• All blood cells have blood groups on outside of
  cell.

• Red Blood Cells contain the protein hemoglobin,
  which carries oxygen

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Introduction to Blood
ABO blood groups found on outside of cell
Hemoglobin picks up and drops off oxygen
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Unknown Stain at a Scene

• Questions to be answered
• Is it blood?
• Is it human blood?
• Whose is it?
• Determine blood type, alcohol content, drugs
  present
• Determine the method(s) in which blood may have
  been deposited

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Forensic Analysis of Blood

1. Visual examination of evidence

1. Presumptive screening test (Is it blood?)

1. Confirmation test (Seriously, is it blood?)

1. Determine species origin (human blood?)

1. Identify the blood (whose blood is it?)

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Forensic Analysis of Blood

1. Visual examination of evidence

1. Presumptive screening test (Is it blood?)

1. Confirmation test (Seriously, is it blood?)

1. Determine species origin (human blood?)

1. Identify the blood (whose blood is it?)

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Presumptive Screening Tests

• Negative result means the questioned stain is not
  likely blood
• Positive result means the questioned stain is
  likely blood

• Presumptive tests produce a color reaction or
  release of light
• Tests rely on catalytic properties of blood
  (hemoglobin presence)
• Three Categories
• Color
• Luminescent
• Fluorescent

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Presumptive Screening Tests

• Color Tests
• Apply chromogen (color changing chemical)
• Apply oxidizing agent (hydrogen peroxide)
• The catalyst of the reaction is hemoglobin

Rapid color change is a positive result. This
means the stain is blood.
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Presumptive Screening Tests

• Color Tests

Remember, other non-blood substances might
catalyze the reaction also.

• Chemical Oxidants
• Plant materials

False Positive A positive result given by a
substance that is not blood.
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Screening Tests

• Based on the peroxidase-like activity of the
  blood
• ?Linked to the hemoglobins heme portion
• Basic reaction
• H2O2 Heme ? 2OH-
• Colorless substrate 2OH- ? Colored product
  2H2O

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Screening Tests

• Extremely Sensitive
• ?Presence of blood detectable even when there is
  no visible staining
• EX phenolphthalein can detect 1 part blood in
  1-10 million parts of dilutent
• Kastle-Meyer Color Test Mixture of
  phenolphthalein and hydrogen peroxide on blood
  cause the mixture to turn pink
• ?Because hemoglobin reacts like a peroxidase
  (enzyme that breaks down hydrogen peroxide),
  reaction will give off oxygen in presence of
  hydrogen peroxide

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Hematest Tablet

• reacts with the heme group in blood causing a
  blue-green color

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TMB

• Tetramethylbenzidine (TMB)
• Positive result Blue-green

• Most common test for blood

1. Rub stain with moist swab
2. Add TMB
3. Add peroxidase

Look for quick blue color
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TMB

• Tetramethylbenzidine (TMB)
• Positive result Blue-green

• Most common test for blood

1. Rub stain with moist swab
2. Add TMB
3. Add peroxidase

Look for quick blue color
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Presumptive Screening Tests

• Chemiluminescence and Fluorescence
• More sensitive than color tests
• May damage blood stain (no blood/DNA typing)

• Used to locate and define areas of blood
• old blood stains
• cleaned floor

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Presumptive Screening Tests

• Luminol
• Method similar to color test
• Luminol is combined with oxidant and sprayed over
  area thought to contain blood
• Emits a blue-white to yellow green glow
• Destroys important markers for DNA fingerprinting

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Presumptive Screening Tests

• Fluorescein
• Fluorescein is combined with oxidant and sprayed
  over area thought to contain blood
• Fluoresces when treated with a UV light
• Fluorescein includes a thickener this makes it
  more effective on vertical surfaces
• Study showed no interference with DNA analysis

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Forensic Analysis of Blood

1. Visual examination of evidence

1. Presumptive screening test (Is it blood?)

1. Confirmation test (Seriously, is it blood?)

1. Determine species origin (human blood?)

1. Identify the blood (whose blood is it?)

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Confirmatory Tests

• Due to the possibility of false positives with
  the presumptive tests, confirmatory tests are
  necessary
• Confirmatory tests involve making crystals that
  detect the presence of hemoglobin
• Teichmann Test and Takayama Test
• Small amount of blood added to microscope slide
• Chemical solution is added
• Slide is heated to form crystals
• Crystals viewed under microscope

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Forensic Analysis of Blood

1. Visual examination of evidence

1. Presumptive screening test (Is it blood?)

1. Confirmation test (Seriously, is it blood?)

1. Determine species origin (human blood?)

1. Identify the blood (whose blood is it?)

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Species Origin

• Most methods test for serum proteins

• Serum proteins are found in all animals, but are
  slightly different

• Species ID methods based on antigen/antibody
  interactions

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Microscopic Views
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Species Origin

• Antigen serum protein

• Antibody produced when foreign serum protein is
  detected

• Certain antibody will only attach to one species
  serum protein

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Species Origin
Antibody is in antiserum
Antigen (serum protein) is in blood sample

• Human antiserum will only attach to human blood
  sample

• Rabbit antiserum will only attach to rabbit blood
  sample

• Dog antiserum will only attach to dog blood sample

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Species Origin

• Ring Precipitin Test

• Blood sample (dilute) in top layer

• Antiserum in heavy bottom layer

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Species Origin

• Ring Precipitin Test

• Blood sample (dilute) in top layer

Precipitate means blood and antiserum species
match

• Antiserum in heavy bottom layer

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Species Origin

• Ring Precipitin Test

Yes line Yes human blood
Human Antiserum
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Species Origin

• Ring Precipitin Test

No line Not human blood
Human Antiserum
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Anti-human test

• Human blood is injected into an animal
• Usually a rabbit
• Animal will produce antibodies against human
  blood
• Serologist will extract antibodies from the
  rabbits serum
• ?Called Human anti-serum

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Anti Human Test

• Layer an extract of the blood stain on top of the
  human anti-serum in a capillary tube
• ?If blood is human, there will be a reaction
  between the human anti-serum and the unknown
  blood and a precipitate will form

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Precipitin test

• An antibody that reacts with its corresponding
  antigen will be used on the unknown stain to see
  if it will precipitate (solidify or crystallize)

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False positives

• Contamination of the blood sample
• Blood is from higher order apes
• Close relatives of humans
• Orangutan
• Chimpanzee
• Gorilla

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Other origin tests

• Gel diffusion
• ?Same basic technique as a precipitin test except
  it is done in a gel plate
• Antigens and antibodies will be attracted towards
  each other
• Electrophoretic method
• ?Apply an electrical charge to accomplish same
  results

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Secretor

• An individual who secretes his or her blood-type
  antigen in body fluids
• Found saliva, semen, vaginal secretions, gastric
  juices
• ?Approximately 85 of people are secretors

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Forensic Analysis of Blood

1. Visual examination of evidence

1. Presumptive screening test (Is it blood?)

1. Confirmation test (Seriously, is it blood?)

1. Determine species origin (human blood?)

1. Identify the blood (whose blood is it?)

42
Genetic Markers in Blood

• If a stain is blood, and it is human blood, then
  whose is it?

• Blood Group Markers

• Protein/Enzyme Markers

• DNA

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Blood Group Markers
Many blood group markers, including

• ABO Markers
• Lewis System
• Rhesus System

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Blood Group Markers

• ABO Markers

• Look at antigens on Red Blood Cells

• In this case, antigens are glycoproteins and are
  attached to the outside of the cell

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Blood Terminology

• ABO blood groups ?based on having an A, B, both
  or no antigens on red blood cells
• Rh factor ?may be present on red blood cells
  positive if present and negative if not
• Antigen ? a substance that can stimulate the body
  to make antibodies. Certain antigens (proteins)
  found in the plasma of the red blood cells
  membrane account for blood type.
• Antibody ? a substance that reacts with an
  antigen
• Agglutination ? clumping of red blood cells will
  result if blood types with different antigens are
  mixed

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Introduction to Blood
ABO blood groups found on outside of cell
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Blood Group Markers

• ABO Markers (antigen and antibodies)

• A person will have antibodies (A or B) to
  whatever blood group he/she doesnt have

• Otherwise, a persons blood would clump up and
  cause death

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Genetics of Blood Types

• Your blood type is established before you are
  BORN, by specific GENES inherited from your
  parents.
• You inherit one gene from your MOTHER and one
  from your FATHER.
• These genes determine your blood type by causing
  proteins called AGGLUTINOGENS to exist on the
  surface of all of your red blood cells.

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What are blood types?
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Blood Group Markers

• ABO Markers

• A cells clump with anti-A
• B cells clump with anti-B
• AB cells clump with both
• O cells do not clump

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Blood Transfusions
A blood transfusion is a procedure in which blood
is given to a patient through an intravenous (IV)
line in one of the blood vessels. Blood
transfusions are done to replace blood lost
during surgery or a serious injury. A transfusion
also may be done if a persons body can't make
blood properly because of an illness.
Who can give you blood? People with TYPE O blood
are called Universal Donors, because they can
give blood to any blood type. People with TYPE
AB blood are called Universal Recipients, because
they can receive any blood type. Rh ? Can
receive or - Rh - ? Can only receive -
Universal Donor
Universal Recipient
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Rh Factors

• Scientists sometimes study Rhesus monkeys to
  learn more about the human anatomy because there
  are certain similarities between the two species.
  While studying Rhesus monkeys, a certain blood
  protein was discovered. This protein is also
  present in the blood of some people. Other
  people, however, do not have the protein.
• The presence of the protein, or lack of it, is
  referred to as the Rh (for Rhesus) factor.
• If your blood does contain the protein, your
  blood is said to be Rh positive (Rh). If your
  blood does not contain the protein, your blood is
  said to be Rh negative (Rh-).

A A-B B-AB AB-O O-
http//www.fi.edu/biosci/blood/rh.html
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Blood Evidence

• Blood samples Can be analyzed to determine
  blood type and DNA, which can be matched to
  possible suspects.
• Blood droplets Can be analyzed to give clues to
  the location of a crime, movement of a victim,
  and type of weapon.
• Blood spatter Can be analyzed to determine
  patterns that give investigators clues to how a
  crime might have happened.

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Blood Group Markers

• ABO Markers (antigen and antibodies)

Blood Type Antigen (blood group) Antibody population
A
B
AB
O
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Blood Group Markers

• ABO Markers (antigen and antibodies)

Blood Type Antigen (blood group) Antibody population
A A Anti-B 40
B
AB
O
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Blood Group Markers

• ABO Markers (antigen and antibodies)

Blood Type Antigen (blood group) Antibody population
A A Anti-B 40
B B Anti-A 10
AB
O
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Blood Group Markers

• ABO Markers (antigen and antibodies)

Blood Type Antigen (blood group) Antibody population
A A Anti-B 40
B B Anti-A 10
AB A B none 5
O
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Blood Group Markers

• ABO Markers (antigen and antibodies)

Blood Type Antigen (blood group) Antibody population
A A Anti-B 40
B B Anti-A 10
AB A B none 5
O H Anti-A Anti-B 45
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How common is your blood type?
46.1
38.8
11.1
3.9
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Blood Group Markers

• ABO Markers

• Testing is similar to species test

• An anti-A, anti-B, or anti-AB antiserum
  (containing antibodies) is reacted with the blood
  stain to detect blood cells

• A, B, or AB blood cells are reacted with a blood
  stain to detect antibodies

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Blood Group Markers

• ABO Markers

• Tests can get complicated with absorbing and
  releasing cells

• Final step is usually testing for agglutination
  (blood clumping)

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Other Markers

• Protein or Enzyme Markers

• Some proteins or enzymes can be in different
  forms (different shapes)

• These differences can be detected by separating
  the proteins in a gel by electrophoresis

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Protein or Enzyme Markers

Ladder
Type 1
Type 2
Type 3

• Charge makes proteins move through gel
• Different shapes move at different rates
• After several minutes, their location will tell
  what type they are.

-
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Protein or Enzyme Markers
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Genetic Markers in Blood

• ABO blood typing and protein analysis may help
  eliminate a suspect.

• Since there are only a small number of types (ABO
  4 types), a match does not mean the stain
  definitely came from a certain person

• DNA testing can identify a person, and is
  becoming just as easy as the above tests