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Identification and Characterization of Blood and Bloodstains
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Summary Introduction to Blood Identification of Blood as Blood
Determining Species Origin of Blood Identifying Humans using Blood
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Introduction to Blood What you should already know:
It’s that red stuff that comes out of your fingers following a kitchen accident.
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Introduction to Blood
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Introduction to Blood
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Introduction to Blood What you should already know:
It’s that red stuff that comes out of your fingers following a kitchen accident. Lets look at blood.
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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.
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Introduction to Blood ABO blood groups found on outside of cell
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Introduction to Blood ABO blood groups found on outside of cell
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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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Hemoglobin picks up and drops off oxygen
Introduction to Blood Hemoglobin picks up and drops off oxygen
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Forensic Analysis of Blood
Visual examination of evidence Presumptive screening test (Is it blood?) Confirmation test (Seriously, is it blood?) Determine species origin (human blood?) Identify the blood (whose blood is it?)
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Forensic Analysis of Blood
Visual examination of evidence Presumptive screening test (Is it blood?) Confirmation test (Seriously, is it blood?) Determine species origin (human blood?) Identify the blood (whose blood is it?)
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Forensic Analysis of Blood
Visual examination of evidence Presumptive screening test (Is it blood?) Confirmation test (Seriously, is it blood?) Determine species origin (human blood?) 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
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Presumptive Screening Tests
Presumptive tests produce a color reaction or release of light Tests rely on catalytic properties of blood (hemoglobin presence)
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Presumptive Screening Tests
Kastle-Meyer Test Apply phenolpthalien (color changing indicator) 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
Kastle-Meyer Test Sample stain with clean cotton swab damp with sterile deionized water Add drop of hydrogen peroxide Add drop of indicator Phenolphthalein Positive result = pink
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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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Presumptive Screening Tests
Other Types of Color Tests Tetramethylbenzidine (TMB) Leucomalachite Green (LMG)
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Presumptive Screening Tests
Tetramethylbenzidine (TMB) Positive result = Blue-green Most common test for blood Rub stain with moist swab Add TMB Add peroxidase Look for quick blue color
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Presumptive Screening Tests
Tetramethylbenzidine (TMB) Positive result = Blue-green Most common test for blood Rub stain with moist swab Add TMB Add peroxidase Look for quick blue color
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Presumptive Screening Tests
Leucomalachite Green (LMG) Positive result = Green Not as sensitive as TMB or specific as phenolphthalein
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Presumptive Screening Tests
Chemiluminescence and Fluorescence Chemiluminescence – light is emitted as a product of the chemical reation Fluorescence – light is emitted when a substance is exposed to a shorter wavelength of light
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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
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Presumptive Screening Tests
Luminol
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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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Presumptive Screening Tests
Fluorescein
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Forensic Analysis of Blood
Visual examination of evidence Presumptive screening test (Is it blood?) Confirmation test (Seriously, is it blood?) Determine species origin (human blood?) 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
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Confirmatory Tests 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
Visual examination of evidence Presumptive screening test (Is it blood?) Confirmation test (Seriously, is it blood?) Determine species origin (human blood?) 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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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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Species Origin Crossed-Over Electrophoresis
Antiserum Blood Stain Gel Holes Antiserum and Blood move toward each other
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Species Origin Crossed-Over Electrophoresis
Antiserum Blood Stain Antiserum and Blood move toward each other If line forms, antiserum and blood match
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Forensic Analysis of Blood
Visual examination of evidence Presumptive screening test (Is it blood?) Confirmation test (Seriously, is it blood?) Determine species origin (human blood?) Identify the blood (whose blood is it?)
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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
Rhesus System
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Blood Group Markers ABO Markers Look at antigens on Red Blood Cells
ABO Blood typing
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Introduction to Blood ABO blood groups found on outside of cell
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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 doesn’t have Otherwise, a persons blood would clump up and cause death
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Blood Group Markers A ABO Markers (antigen and antibodies) B AB O
Blood Type Antigen (blood group) Antibody % population A B AB O
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Blood Group Markers ABO Markers (antigen and antibodies) A Anti-B 40 B
Blood Type Antigen (blood group) Antibody % population A Anti-B 40 B AB O
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Blood Group Markers ABO Markers (antigen and antibodies) A Anti-B 40 B
Blood Type Antigen (blood group) Antibody % population A Anti-B 40 B Anti-A 10 AB O
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Blood Group Markers ABO Markers (antigen and antibodies) A Anti-B 40 B
Blood Type Antigen (blood group) Antibody % population A Anti-B 40 B Anti-A 10 AB A & B none 5 O
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Blood Group Markers ABO Markers (antigen and antibodies) A Anti-B 40 B
Blood Type Antigen (blood group) Antibody % population A Anti-B 40 B Anti-A 10 AB A & B none 5 O H Anti-A & Anti-B 45
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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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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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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, but still expensive and time consuming.
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