T. Trimpe 2006 http://sciencespot.net/

Objectives

You will understand:That an antibody and an

antigen of different types will agglutinate, or clump, when mixed together.

That the significance of the evidence depends on a characteristic’s relative occurrence in the population.

Objectives, continued

You will be able to:Determine the blood type of a

simulated bloodstain using the ABO/Rh system.

Explore bloodstain patterns as a function of velocity, direction, and height of fall.

Determine the genetics behind ABO/Rh antigens

Determine whether a stain is blood.

Serology

Serology is the examination and analysis of body fluids. A forensic serologist may analyze a variety of body fluids including saliva, semen, urine, and blood. From 1950 to the late 1980s, forensic serology was a most important part of lab procedures. With the development of DNA techniques, more time, money, and significance were placed on developing DNA labs. However, with limited funds and the time required for DNA testing, most labs still use many of the basic serology testing procedures.

Historical Perspective of Blood TypingAround 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 percent of Caucasians, 94 percent of African Americans, and 99 percent of all Asians Americans are Rh positive.

Blood TerminologyABO blood groups—based on having 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 cell’s 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

What makes up our blood? RED BLOOD CELLS (erythrocytes) – The most

abundant cells in our blood; they are produced in the bone marrow and contain a protein called hemoglobin that carries oxygen to our cells.

What makes up our blood?

PLATELETS (thrombocytes) – The clotting factors that are carried in the plasma; they clot together in a process called coagulation to seal a wound and prevent a loss of blood

What makes up our blood?

PLASMA – The yellowish liquid portion of blood that contains electrolytes, nutrients and vitamins, hormones, clotting factors, and proteins such as antibodies to fight infection.

What makes up our blood? White Blood Cells (leukocytes) – they are

part of the immune system and destroy infectious agents called pathogens

Blood Facts

The average adult has about FIVE liters of blood inside of their body, which makes up 7-8% of their body weight.

http://www.bloodbankofalaska.org/about_blood/index.html

Blood Facts

This red liquid 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 fights against infection and helps heal wounds, so we can stay healthy.

Blood Facts

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.

How is our blood type determined?

Your blood type is established before you are BORN, by specific GENES inherited from your parents.

You receive one gene from your MOTHER and one from your FATHER.

More to come later……

You blood type These two genes determine your blood type

by causing proteins called AGGLUTINOGENS to exist on the surface of all of your red blood cells.

http://www.lanecountyblood.org/images/other/bloodfacts.jpg

45%

40%

11%

4%

How common are the 4 blood types?

BLOOD GENETICS

Blood Genetics

The human ABO gene is on chromosome 9.

Everyone has two copies of chromosome 9 so you have two ABO genes.

One copy is inherited from our mother, the other from our father.

Alleles

There are three versions (called “alleles”) of this blood type gene: A, B, and O.

A person’s blood type is determined by which allele he/she inherits from each parent.

Pheno vs. Geno

The genetic makeup of an organism is called the “genotype”.

The “phenotype” is the visible properties of an organism.

In this case, the A, B, and O allele combination a person has is their genotype

Their blood type is their phenotype.

Dominant vs. Recessive Genes The “A” allele is dominant and so is the

“B” allele. Together though, the “A” and “B” alleles

are co-dominant. The “O” allele is recessive.

Determining the Genotype

The blood type gene has three different alleles:

IAIA is Type A IAIB is Type AB

IBIB results in Type B ii is Type O

Determining the Genotype Scenario:

Mom has the alleles IAIA for blood type and Dad has the alleles IBIB blood type.

What will be the blood type for their child?

Punnett Sqaures

IA IA

IB

IB

Practice

Suppose that a mother has blood Type A and genotype IAi and the father has blood Type B and genotype IBi. Draw a Punnett square to show the possible genotypes of their children.

What are the phenotypes of the kids?

Practice

IB i

IA

i

Blood Types

The alleles we discussed “code” for blood type.

What they REALLY “code” for is a specific enzyme.

That enzyme creates specific antigens on your RBC.

Reminder….

An antigen is a protein (encoded from the right enzyme) that “sits” on the surface of your RBC.

There are 2 different blood antigens, A and B.

If you have the A antigen, you have type A blood.

If you have the B antigen, you have type B blood.

Antigens of the Surface of the RBC

Antibodies

Blood plasma is packed with proteins called antibodies.

The body produces a wide variety of antibodies that will recognize and attack foreign molecules.

A person’s plasma does not contain any antibodies that will bind to molecules that are part of his or her own body.

Blood Transfusions

It is important to carefully match the donor and recipient blood types.

If the donor’s blood cells have antigen that are different from those of the recipient, antibodies in the recipient’s blood recognize the donor blood as foreign.

This triggers an immune response resulting in blood clotting.

Blood Transfusions

Antigen Antibody Can donate to…

Can receive from…

A

B

AB

O

Relative Abundance of Blood Types

A B AB O

40-42% 10-12% 3-5% 43-45%

Interesting Facts

Men generally have more red blood

cells than women.    Rare blood types exist in addition to

the basic ABO system.

Rhesus Factor (Rh)

If a person has a positive Rh factor, this means that their blood contains a protein that is also found in Rhesus monkeys. 

Most people (about 85%) have a positive Rh factor

Rh is expressed as either positive or negative. 

The Rh factor, like other antigens, is found on the surface of the red blood cells. 

Stats

 O+ 1 in 3 persons O- 1 in 15 persons A+ 1 in 3 persons A- 1 in 16 persons B+ 1 in 12 persons B- 1 in 67 persons

AB+ 1 in 29 persons AB- 1 in 167 persons

Can Blood Be Individualized?

Is based on the typing of proteins and enzymes. Blood proteins have subtypes.

Most people are familiar with at least one common polymorphism in blood: Hb, which causes sickle-cell anemia.

You can also extract DNA from it.

MNS Blood System Since blood typing is useful in forensic

because it can show that two samples did not have the same origin, this exclude potential suspects. One would want to take into account as many factors as possible. The simplest way would be to exclude as many people as possible with taking into account as many surface antigens as possible.

M, N and S are three additional surface antigens that can be used in cases such as paternity.

MNS Typing

Type of Blood Percentage of Population

M 30

N 20

S 48

Number of people with a blood type 3 surface antigens as probability

1. Determine % of people with that surface antigen

2. Determine the ratio by dividing 100 by the %

3. Multiple the ratios together

4. Obtain the # of people out of 100 with those 3 antigens

DATA Type of Blood % of People

A 41

B 10

AB 4

O 45

Rh + 85

Rh - 15

M 30

N 22

S 48

Determine the likelihood… 1. Of an individual with Type A, N and Rh –

2. Of an individual with Type AB, S and Rh +

3. Of an individual with Type O, M, Rh -

Blood Typing

Blood type A has antigen A on the surface of the cell and will agglutinate with blood type B.

Blood type B has antigen B on the surface of the cell and will agglutinate with blood type A.

Blood type AB has antigens A and B on the surface of the cells and will not agglutinate with either type A or type B blood.

Blood type O has neither antigen A nor B and will not agglutinate.

Blood Groups

Type Antigen Antibody Can GiveBlood To

Can GetBlood From

A

B

AB

O

A

B

A and B

NeitherA nor B

B

A

NeitherA nor B

A and B

A, AB O, A

B, AB O, B

AB

A, B, O, AB

A, B, O, AB

O

What are blood types?

http://learn.genetics.utah.edu/units/basics/blood/types.cfm

There are 3 alleles or genes for blood type: A, B, & O. Since we have 2 genes, there are 6 possible combinations.

Blood Types

AA or AO = Type ABB or BO = Type B

OO = Type OAB = Type AB

Population Distribution of Blood Types in the U.S.

Type Percent

O

A

B

AB

45

40

11

4

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.

http://www.fi.edu/biosci/blood/rh.html

Rh Factors 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-

How can blood be used as evidence?

Blood samples – Can be analyzed to determine BLOOD TYPE and DNA, which can be matched to possible suspects.

How can blood be used as evidence?

Blood droplets – Can be analyzed to give clues to the location of a WOUND, movement of a VICTIM, and type of INJURY.

Blood Typing Use the results shown to determine the blood type

for each sample.

- = No Clumps

+ = Clump

A

B

Rh

Blood Type: ___

To determine a person’s blood type, we add three different serums to blood samples and see if clumps form.

Clumps = Positive (+)No Clumps = Negative (-)

What would the blood type be for this sample?

A

B

Rh

Blood Type: ___

A

B

Rh

Blood Type: ___

A

B

Rh

Blood Type: ___

A

B

Rh

Blood Type: ___

A

B

Rh

Blood Type: ___

Blood Typing Use the results shown to determine the blood type for each sample.

- = absent

+ = present

Bloodstain Pattern Analysis

How can blood be used as evidence?

Blood spatter – Can be analyzed to determine PATTERNS that give investigators clues to how a crime might have happened.

Objectives of Bloodstain Pattern Analysis

Can provide information about:Origin of bloodstainObjects that produced bloodstainDistance from bloodstain to targetPosition of victim and assailantMovement of victim and assailantNumber of blows/shots

Physical Properties of Blood Viscosity

Thickness of a liquid Surface tension

Adhesion○ Attractive forces between unlike molecules

Cohesion○ Attractive forces between like molecules

Capillary action○ When surface tension causes a liquid to be drawn

up in opposition to gravity

Size of Bloodstain

Shape of BloodstainOn impact with hard, smooth, nonporous surfaces (tile, glass)

○ Little to no distortion of stain○ Surface tension of blood drop resisting rupture

On impact with rough/textured, porous surfaces (concrete) Disrupts surface tension of blood drop

• Stain will exhibit distortion, irregular shapes, and spiny edges

– Spines = pointed edges of bloodstains that radiate away from the center of the stain

• May also see satellite spatter

Angle of Impact

A spherical drop of blood impacting a horizontal surface should create a circular stain

A drop of blood impacting a nonhorizontal surface creates a more elongated or elliptical stain due to the blood “skidding” across the target surfaceThis allows you to make 2 determinations

○ Directionality of stain○ Angle of impact

Directionality

Directionality

Parent Stain with tail pointing

in direction of travel

Tail of cast off stain points

back to parent stain

Angle of Impact

The more acute the angle of impact = the greater the elongation of the stain Length increases and width decreases

By accurately measuring the length and width of the bloodstain then, the angle of impact can be calculated using the following equation:

Angle of Impact = arc sin (width/length)

Angle of Impact

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

Points of Convergence

Points of Convergence

Bloodstain Categories 3 Categories

Passive○ Only acted on by gravity and friction

Spatter○ Subject to some force other than gravity and friction

Altered○ Stains whose physical appearance has been changed in

some way

Passive Bloodstains Includes:

Transfer stains/swipesFree flowing blood and dropsLarge volume blood loss

○ Saturation, pools and splashes

GRAPHIC PICTUREs FOLLOWING 3 SLIDEs

Passive Bloodstains

Passive Bloodstains

Passive Bloodstains

Spatter Bloodstains

Includes:Satellite spatterImpact mechanisms

○ Gunshot ○ Beating○ Stabbing

Projection mechanisms○ Cast-off○ Arterial spurting○ Expired blood

Classification of Spatter Bloodstains

Low-Velocity Impact Spatter Less than 5 ft/sec Blood spots 4-8 mm in diameter

Medium-Velocity Impact Spatter– 5-100 ft/sec– Blood spots 4 mm in diameter or less

High-Velocity Impact Spatter– Over 100 ft/sec– Blood spots 1 mm in diameter or less

Spatter Bloodstains

Altered Stains Includes:

Blood decompositionClottingDiluted bloodDiffused bloodRemoved blood

Altered Bloodstains

Diffusion

Bloodstain Categories

ID THE STAIN

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

OVERVIEW Presumptive Tests for Blood Determination

Kastle-Meyer color test—a mixture of phenolphthalein and hydrogen peroxide; the hemoglobin will cause the formation of a deep pink color if blood is present

Hematest tablet—reacts with the heme group in blood, causing a blue-green color

Luminol test—reaction with blood to produce light

Color tests to identify human blood

The Benzidine color test was used for years to identify human blood

But benzidine was identified as a carcinogen, so nowadays the phenolphthalein test is performed instead.

The phenolphthalein test is also known as the Kastle-Meyer color test

Both, the benzidine and the Kastle-Meyer test look for the presence of hemoglobin in the sample.

The Kastle-Meyer Test

When blood, phenolphthalein and hydrogen peroxide are mixed, the hemoglobin in the blood will cause the normally colorless phenolphthalein to a bright pink color

This test can yield some false positives Certain vegetable matter can produce

the bright pink positive (such as potatoes)

Death By Potato? Although the Kastle-

Meyer test can give a false positive in the presence of certain plant matter, it is unlikely that this plant matter would be present at a crime scene and be mistaken for blood.

Other blood detection tests

Doctors use Hemastix© strips to detect blood in urine

But these strips can be used at a crime scene to detect fresh or dried blood.

One simply moistens them with distilled water and wipes the bloodstain

A color change to green is a positive indicator of blood

Bayer Hemastix©

Luminol Luminol is a chemical that exhibits

chemiluminescence, with a striking blue glow, when mixed with an appropriate oxidizing agent. It is a white to slightly yellow crystalline solid that is soluble in water and most polar organic solvents.

Usually, a solution of hydrogen peroxide (H2O2) and a hydroxide in water is used as the activator.

In the presence of a catalyst such as an iron compound, the hydrogen peroxide is decomposed to form oxygen and water:

What do you see with luminol?

It glows a bright blue in the dark, when it comes in contact with blood

Advantages of using Luminol Allows one to detect stains that would

not be ordinarily be visible Extremely sensitive - can use it in very

dilute concentrations This allows the CSI to spray large areas

with it It does not interfere with DNA, so a CSI

can collect samples for DNA analysis even after it was sprayed with luminol.

Bloody Footprints that were wiped

Drawbacks

Luminol glows even in the presence of certain other fluids – semen, feces, bleach, tonic water, etc.

Luminol glows in the presence of certain vegetable matter - Mr. Potato head strikes again!

Thanks to Quinine

Crystal Tests

Less sensitive than color tests They involve the crystallization of certain

components of blood when certain chemicals are added

Takayama test and Teichman test Not used as much by CSI

So we know it’s blood – but is it human? The Precipitin Test

Rabbits injected with human bloodThey make antibodies in their

serumThey are bled and the serum

recoveredThe serum is called human

antiserum, because it will react to human antigens

It will cause coagulation when mixed with human blood

Precipitin Test

(Human blood)

(Human antiserum made in rabbits)

Gel Diffusion Test Antigen (blood) and antibodies (human

antiserum from rabbits) are placed in separate wells in a gel.

They are forced to move towards each other.

If they bind to each other, they form a single line of precipitate in between the wells

This is a positive for human blood

DNA TEST

DNA testing is the ultimate It has made most other tests for

determining if the blood is human, obsolete

Analyzing Bloodstain Patterns1. Surface Texture

The harder and less porous the a surface, the less spatter results

2. Direction of travel The pointed end of a bloodstain always faces the

direction of travel3. Impact angle

If the strike angle is 90deg then the stain is circular. As the strike angle increases, the stain becomes more and more elongated

4. Origin of blood spatter If you draw straight lines through the long axis of

several blood stains, the point where the lines converge is the origin of the blood spatter