Forensic Serology

Serology-study of biological fluids involving specific antigen and serum antibody

reactions

• 1901-Karl Landsteiner-ABO blood type system• Blood types based on presence of

agglutinogens (agglutinating antigens) on surface of red blood cells (erythrocytes)

• Proteins that stimulate production of antibodies

• On surface of erythrocytes• Types- A and B• Antibodies-produced against antigens not

present• Ex. Type A blood has A antigens, but B

antibodies

Antigens

Blood Types

Blood Type

Antigen Antibodies in plasma

Can give to

Can receive from

A A Anti B A,AB O, A

B B Anti A B,AB O.B

AB AB No A or B AB O,A,B,AB

O Neither A nor B

A and B O,A,B,AB

O

http://nobelprize.org/educational_games/medicine/landsteiner/

Frequency of Blood Types

Blood Factors

• If mixed-blood agglutinates (solidifies) -death

• Rh factor (antigen D)• More than 100 blood factors

Blood

1. Cells-Red blood cells

(Erythrocytes)Platelets (Thrombocytes)White blood cells

(Leukocytes)

2. Enzymes3. Proteins4. Fluid5. Inorganic

substances (gases)

Blood Plasma

• Liquid part of blood • Serum (liquid that separates from blood)

and proteins in water• 55% of blood volume• Fibrin-blood clotting protein

Erythrocytes

• Red blood cells• Transport oxygen and carbon

dioxide• Has hemoglobin-blood protein

carries oxygen in blood• Most numerous blood cells

Leukocytes (White Blood Cells)

• Monocytes• Lymphocytes• Neutrophils• Part of Immune System• Fight infections

Platelets• Thrombocytes• Involved in blood clotting

Antiserum

• Serum that contains antibodies (proteins that destroy specific antigens ;immunoglobins)

• Used in determining blood types• If antiserum causes agglutination,

then the blood type can be determined

Bloodstain Characteristics

• As blood dries-harder to type• Erythrocytes lyse, or break open, upon

drying• No cells in dried bloodstain-antigens will

be present

• Determine if it is blood• Determine species• If human blood-determine if it can be

individualized• Blood is class evidence

Guideline for Examining Blood

1. Is it blood?2. From what species did the blood originate?3. If the blood is human, how closely can it be matched to one individual?

Forensic Characterizations of Bloodstains

Is it Blood?Kastle-Meyer Color Test

• Phenolphthalein• Hemoglobin-peroxidase-like activity• Peroxidases-speed up oxidation of

compounds by peroxides• Blood, phenolphthalein, and hydrogen

peroxide-hemoglobin is deep pink

1.) Hydrogen peroxide2.) Heme has peroxidase-like activity-breaks up

peroxides.H2O2 decomposed to H2O and oxygen3.) Oxygen combine with the KM, phenolphthalein color-

indicator4.) When KM solution is prepared-boiled remove oxygen-

When it turns colorless-shows KM had most of its oxygen removed and is now "oxygen hungry". So... now we have the makings of a kind of love story: the KM wants oxygen, and there's a lonely oxygen radical floating around, un-spoken for

5.) Pink color: occurs when oxygen radical "hooks up" with KM reagent-oxidation of KM and causes it to turn pink.

Luminol TestChemiluminescence of luminolThe most sensitive chemical test that is capable of presumptively detecting bloodstains diluted up to 300,000 times

• Luminol (C8H7O3N3) mixed with hydrogen peroxide and hydroxide

• If hemoglobin and luminol interact-iron in hemoglobin catalyzes rxn b/w H2O2 and luminol

• Redox rxn-luminol loses N and H and gains O-produce 3-aminophthalate

• 3-aminophthalate in energized state –e-s in oxygen are boosted to higher orbitals-e-s then fall back to lower energy level-emit extra energy as light photons

• http://www.howstuffworks.com/framed.htm?parent=luminol.htm&url=http://www.shsu.edu/%7Echm_tgc/JPPdir/JPP1999/

Immunoassay TechniquesPresumptive test for detection of drugs in blood and urine

Antibodies reacting with drugs don’t occur naturally

Produced in animals by combining drug with a protein and injecting into animal

Drug-protein complex acts as antigen stimulating animal to produce antibodies

The recovered blood serum of animal contains specific antibodies to the drug-This is human antiserum

EMIT- Enzyme Multiplied Immunoassay Technique-drug screening in urine and blood-add antibodies to blood or urine, then add chemically labeled drug- will compete with drugs in urine or blood (if present) for antibody binding sites. The number of remaining unbound labeled drug-proportional to drug’s concentration in urine.ex. Methadone-add methadone antibodies and chemically-labeled methadone to urine- ex. THC tetrahydrocannabinol- metabolite-THC-9-carboxylic acid-antibodies are prepared against this metabolite-one-millionth of a gram-can detect up to 10 days after use

Steps required to produce monoclonal antibodies

Polyclonal vs. Monoclonal Antibodies

• Polyclonal antibodies-series of antibodies which can attack some site on antigen-vary in specificity and binding abilities

• Monoclonal antibodies-identical antibodies that interact with one site on an antigen

Monoclonal antibodies

• Immunoassay tests for abused drugs made with monoclonal antibodies

Precipitin Test

• Test to determine species of blood• Precipitin-an antibody that reats with its

corresponding antigen to form a precipitate

• Animal blood injected with human blood-antibodies form that react with human blood to neutralize its presence

• Antibodies can be recovered in serum• Serum is human anti-serum

•Precipitin tests•Blood is placed on antiserum in tube•Blood will react with anitbodies in antiserum•Leaves precipitin band

Gel Diffusion

Blood Enzymes and Proteins

• Enzymes-proteins that regulate chemical reactions

• Polymorphic enzymes-enzymes of many forms

• Ex. PGM-Phosphoglucomutase-ten types of PGM

Secretors

• 80 % of individuals are secretors• Secretors-antigens in most body fluids

(saliva, semen) and blood

Bloodstains and Probability

• Antigens, enzymes, and proteins occur independently

• Probability-product of their distributions-Rule of Multiplication

• Ex. If bloodstain is type A, it originated from 42% of population. If it has PGM 1-origin narrowed to 24%

• 42% X 58%

Study performed by Herbert MacDonell:

1. Surface texture- harder and less porous surface-less spatter

2. When blood drop hits hard surface-breaks up on impact-casting off smaller droplets

3. Impact angle of blood-Inverse Sin W/LW=width of stainL=length of stain

Area of convergence on a two-dimensional plane. Convergence represents the area from which the stains emanated.

http://www.nfstc.org/links/animations/images/blood%20spatters.swf

Categories of BloodstainsPASSIVE

                                                     

TRANSFER

                                                                                   

PROJECTED

                                                                                            

                       

By measuring the length and width of a bloodstain, the impact angle can be calculated

                                                                     SIN < = Width (a) 1.5cm Length (c) 3.0cm Width (a) 1.5cm = SIN < Length (c) 3.0cm

0.5 = SIN <

< = 30 degrees

Collection of Wet Blood

• Collected and refrigerated• Insufficient amount-air-dry • No plastic containers• Each stained article-packaged in paper

bag or cardboard box

Drying Cabinets in Crime Lab

Collection of Dried Bloodstains

• Stained sample packaged• Sample of unstained surface removed-

substrate control to confirm test results from stain and not substrate on which it was deposited

• Absorb blood onto damp swab

Blood Collection

• Bloodstains compared to blood specimens from victims and suspects

• Collect 5cc from individuals-add anticoagulant EDTA (ethylenediamine tetraacetic acid)-inhibits enzymes from degrading DNA

Heredity of Blood Types

• Genes-basic unit of heredity-located in DNA

• Chromosomes-structures made of DNA• Humans-46 chromosomes in somatic cells• 23 chromosomes in gametes• Zygote-fertilized egg• Alleles-forms of genes

Multiple Alleles of Blood Types

• Homozygous-gene pair made of two similar genes-ex. AA and OO

• Heterozygous-gene pair made up of two different genes-AO

• A and B-dominant genes• O-recessive gene• AB-codominant

Genotype and Phenotype• Genotype-pair of alleles-no laboratory test to

determine genotype ex. AA or AO• Phenotype-physical characteristic-ex. Type A

blood• Punnett Squares used to determine genotypes• Used in Paternity Testing

Mother

Fathero

A

B

o

Forensic Characterization of Semen

• Each Male Ejaculate-2.5-6 mL of seminal fluid

• Each mL-100 million or more spermatozoa

• Acid-Phosphatase Color Test-enzyme secreted by prostate gland

• Oligospermia-low sperm count• Aspermia-no spermatozoa in seminal fluid

• If semen is detected, but contains no spermatozoa, how do you prove presence of semen?

• Protein called p30-unique to seminal plasma