Hematology

What is Karl Landsteiner credited with discovering?

Blood Compositio

n

Key Terms

• Anticoagulant: an agent that prevents the clotting of blood. – Examples are EDTA, Citrate and Heparin

• Capillary: small blood vessel that connects arterioles and venules

• Hematoma: a subcutaneous mass of blood at a venipuncture site

Key Terms

• Hemoglobin: the oxygen carrying molecule of red blood cells

• Hemolysis: the breakdown of red blood cells, with the release of hemoglobin into the plasma or serum. Cannot use hemolyzed samples in lab tests

• Icteric: jaundiced; dark yellow or greenish serum or plasma

Key Terms• Lipemic: having abnormally high level

of fat. Milky looking samples

• Plasma: pale yellow part of whole blood; contains all clotting factors

• Serum: liquid portion of blood without the protein fibrinogen, which is one of the clotting factors; clot removed

Key Terms

• Agglutination: the clumping together of rbc by the action of an antibody– When A cells are added to a blood sample,

agglutination will occur only in the presence of anti-A

• Serology: the study of antigen – antibody reactions using laboratory tests

Function of Blood

• Transporting fluids such as:– Nutrients from digestive tract

– O2 from lungs

– Waste from cells– Hormones

• Aids in heat distribution• Regulates acid-base balance

Composition of Blood

• Plasma: liquid portion of blood w/out cells– Contains all of the following

• Water Nutrients• Electrolytes Metabolic waste

product• Hormones Vitamins and

enzymes• Plasma proteins such as fibrinogen, albumin and

globulin

Composition of Blood: Erythrocytes

• Red blood cells are responsible for:– Transport of oxygen and nutrients

– Removal of waste and CO2 from the cells

– Distribution of heat

• Hemoglobin: the O2 carrying potential

Composition of Blood: Leukocytes

• WBC are responsible for:– Phagocytosis – to engulf and absorb waste

material and harmful microorganisms in the blood stream and tissues

– Synthesis of antibody molecules– Inflammation process– Production of heparin – component found in

lung and liver tissue which have the ability to prevent clotting of blood.

• Heparin used in the treatment of thrombosis

Cell Morphology

Lymphocyte Segmented Neutrophil

Platelet

rbc

Composition of Blood: Leukocytes

• Types of Leukocytes– Granulocytes

• Neutrophils• Eosinophils• Basophils

– Agranulocytes• Lymphocytes• Monocytes

Composition of Blood: Thrombocytes

• Platelets – the smallest of the solid components of the blood

• Responsible for the clotting process

• Coagulation: term for clotting

• Embolism: a blood clot which is moving through the body

Forensic Characterization of Blood

• 1st questions a criminalist has to answer is:– Is it blood– If yes, is it human– If yes, can it be associated w/ a particular person

• Preliminary color test for blood is the Benzidine color test but this is carcinogenic so phenolphthalein is used and is known as the Kastle-Meyer Color test

Forensic Characterization of Blood

• Hemoglobin possesses peroxidase like activity which when mixed with phenolphthalein and hydrogen peroxide it will cause the formation of a deep pink color

• Kastle-Meyer is not specific for blood as some vegetables such as potatoes and horseradish contain peroxidase and can react; however they should not be common at a crime scene so it is often considered a good indicator

Forensic Characterization of Blood

• Hemastix strips can also be used to detect the presence of blood

• Luminol test is another presumptive test for blood which produces light rather than a color reaction. Objects being tested must be in a dark location to view the luminescence (emission of light).– Extremely sensitive and can detect blood

diluted up to 300,000 times– Luminol will not interfere with other DNA testing

Forensic Characterization of Blood

• Once blood is found it must be determined to be human– Precipitin test is the standard test– Reagents are available to determine if blood is

dog, cat or deer– A positive test is a cloudy ring or band at the

point where the two liquids meet– Only a small amount of sample is required– Precipitin test is very sensitive and can test

positive on a sample as old as 10 years– Extracted tissue samples from mummies as old

as 4,000 years have tested positive

Forensic Characterization of Blood

• Gel diffusion is another method which can be used.

• Antigen and antibody assay

• Gel electrophoresis can also be used to determine if a sample is human blood or not

• Antigen and antibody assay• Uses electrical current and ppt. line forms where the

two samples meet if it is a positive reaction

Blood Types

• Four Major Groups– A B AB O

• Blood types are inherited from your parents

• Antigen is present on the red blood cell; typing is done w/rbc

• Antibody is present in the plasma; antibody screening done on plasma

Blood Types

• O negative– Universal donor– It carries no antigen

• AB positive– Universal recipient– It carries no antibodies in the plasma

• 43% of population are O, 42% A, 12% B and 3% AB

Rh Factor (D antigen): found on the surface of

rbc• Rhesus factor:

discovered in rhesus monkeys in 1937

• Can be phenotypically positive or negative

• Positive is dominant over negative– If positive is present,

then you will express positive phenotype

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+- --

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+ -

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+

Blood Types: ABO

• Controlled by GENETICS!!!!!

• Two Categories– ABO and Rh

AA AO

AO OO

A O

A

O

AA AB

AB BB

AB BO

AO OO

A O

B

O

A B

A

B

Blood Types: What Ag do they have?

• A positive blood has which Ag present on rbc?– A antigen and Rh antigen

• B negative blood has which Ag present on rbc?– B antigen

• AB + blood type has which Ag present?– Both A and B Ag and Rh antigen on the rbc

Who can donate to whom?

• A + donor: what blood types can this pt. receive?– A+, A-, O+, O-

• Your blood type is B-, what blood types can donate to you?– B- and O- AB- as a last resort

ABO Blood Types

Agglutination Reactions

Anti-A Anti- B Anti-A & Anti B

A Yes No Yes

B No Yes Yes

AB Yes Yes Yes

O No No No

Examples of ABO blood typing

O negative carries no Ag and therefore does not react with any Anti A, B, AB

Neg reaction

Pos reaction

Anti- Rh

Rh + YES

Rh - NO

Rh Antigen and Antibody Interaction

Normal Ranges

• RBC: female 3.6-5.0x106mm3 male 4.2-5.4x106mm3

• WBC: 4.5-10.5x103mm3 (African Americans is sltly lower 3.2 is still normal)

• HCT: female 36-48% male 42-52%

• Hgb: female 12-16 g/dL male 14-17.4 g/dL

• Platelets: 140-400x103mm3

Blood Cell Maturation

Blood Splatter Analysis

• Location, distribution, and appearance of blood stains are an important part of forensics

• Investigators try to determine:– Direction– Dropping distance– Angle of impact

• Splatter analysis is often used for crime scene reconstruction

Blood Splatter Analysis

• Factors which influence stain patterns are:– Surface texture– Direction of travel

• Pointed end of bloodstain always faces its direction of travel

– Angle of impact is determined by measuring the degree of circular distortion of the stain

• Blood striking a surface at right angles gives rise to a nearly circular stain

• As the angle decreases, the stain becomes elongated in shape

DNA

• DeoxyriboNucleic Acid• Found in the nucleus• 46 chromosomes• 25,000 genes• Structure determined by Crick and

Watson• DNA fingerprinting by Alec Jeffreys

Structure of DNA

• A polymer made of repeating nucleotides

• Nucleotide consists of a deoxyribose sugar, a phosphate, and a nitrogen base (Adenine, Thymine, Cytosine, or Guanine)

• Double stranded, helical• Complementary base pairing, A=T, G=C

DNA at Work

• DNA in nucleus is copied into a strand of RNA (transcription)

• RNA is read at the ribosome to make assemble amino acids into proteins (translation)

• Every 3 bases on DNA codes for a different amino acid

Replication of DNA

• Replication – the synthesis of new DNA from existing DNA in the nucleus

• DNA polymerase assembles new DNA strand and proof- reads it• Replication occurs in nucleus prior to

cell division

Polymerase Chain Reaction

• A technique for replication, or amplifying, a portion of DNA outside

the cell• Each cycle doubles the number of copies• 1 1x107 in 30 cycles

DNA Typing with Tandem Repeats

• Region of chromosome that contains multiple copies of a core DNA sequence arranging in a repeating fashion between the coding regions (genes)

• Restriction Fragment Length Polymorphisms used enzymes to cut the DNA around these tandem repeat sites and then run them on a gel electrophoresis

• A Southern blot was then performed and radioactive probes were hybridized to help visualize the RFLPs

RFLPs

PCR

PCR has the following advantages: 1. PCR can use shorter sequences 2. shorter pieces more stable 3. smaller amounts of DNA can be used (10-9 gram)

Short Tandem Repeats (STRs)

• A region of a DNA molecule that contains short segments of 3-7 repeating base pairs.

• Generally less than 450 bp long• Less degradation• Can be PCR’d• Can multiplex a large number of

these STRs at once• US uses 13 STRs for tests

• Capillary electrophoresis• Sex Identification by focusing on the amelogenin gene