Chapter 8 Forensic Serology

1. Chapter 8 Forensic Serology Courtesy of C. Fanning

2. Unit Objectives for Serology • Understand the anatomy and philology of blood. Understand the concept of the antigen – antibody interactions and how they are applied to species identification and drug identification. • Contrast chromosomes and genes including Punnett squares, genotypes and phenotypes of offspring. • List and understand how whole blood is typed. • List and describe forensic tests used to characterize a stain as blood. • Describe the proper collection of physical evidence in a rape case.

3. Nature of Blood • The word blood refers to a highly complex mixture of cells, enzymes, proteins, and inorganic substances. • Plasma, which is the fluid portion of blood, is composed principally of water. • Red blood cells (erythrocytes), white blood cells (leukocytes), and platelets are the solid materials suspended in plasma. • Antigens, usually proteins, are located on the surface of red blood cells and are responsible for blood-type characteristics.

4. Nature of Blood

5. The Circulatory System • The main arteries carry blood to the body. • Veins carry blood back to the heart.

7. Arteries and Veins • Arteries carry blood away from the heart and veins return blood to the heart. Veinsare generally larger in diameter, carry more blood volume and have thinner walls in proportion to their lumen. Arteries are smaller, have thicker walls in proportion to their lumen and carry blood under higher pressure than veins.

8. The Circulatory System • Blood spirts out from the arties and veins because of blood pressure provided by the heart pumping the fluid around.

9. Blood Cells • Blood cells are formed in the bone marrow. All blood cells arise from the same bone marrow stem cells. Stem cells are immortal, meaning they never die (at least not until you do). • Stem cells are also undifferentiated. meaning they have not yet developed into a particular cell type. Furthermore, stem cells are pluripotent, meaning they have the potential to become any type of blood cell. These immortal, undifferentiated, pluripotent stem cells give rise to erythrocytes, leukocytes and platelets.

10. Leukocytes: Types and names • The diagram below illustrates the different types of blood cells. Leukocytes, also known as white blood cells, are a group of related cell types that involved in immune function. • Leukocytes include neutrophils, eosinophils, basophils, lymphocytes and monocytes.

11. Leucocytes: White Blood Cells and Erythrocytes : Red Cells

12. Erythrocytes: Red Cells • Erythrocytes, also known as red blood cells (RBCs), function to transport oxygen in the blood. The shape of erythrocytes is ideal for this function, they are biconcaved discs. • This shape increases the surface area-to-volume ratio of the cell, thus increasing the efficiency of diffusion of oxygen and carbon dioxide into and out of the cell. They have a flexible plasma membrane. This feature allows erythrocytes, which have a 7mm diameter, to squeeze through capillaries as small as 3 mm wide.

13. Red Blood Cells – cont. • Erythrocytes contain tremendous amounts of hemoglobin, the protein that binds oxygen. In order to make room for more hemoglobin to carry more oxygen, erythrocytes loose their nucleus and other organelles as they develop in the bone marrow. Because they lack a nucleus and other cellular machinery, erythrocytes cannot repair themselves when damaged, consequently they have a limited life span of about 120 days.

14. Erytropoesis: Production of Blood Cells • The removal of old and dying erythrocytes is carried out by the spleen. Erythrocytes, which represent the most numerous cell type in the body die at a rapid rate, 2-3 million erythrocytes die every second. Erythrocyte production must equal erythrocyte death or the cell population would decline. Erythrocytes are produced through a process called erythropoesis.

15. Hematocrit • Whole blood is composed of plasma (liquid), cells and platelets. If whole blood is placed into a tube and centrifuged, the cells and the plasma will separate. • The erythrocytes will pack into the bottom of the tube, the plasma will be at the top of the tube, and the leukocytes and platelets will form a thin layer in the middle. • The hematocrit is defined as the percentage of whole blood made up of erythrocytes.

16. Hematocrits vary by Gender • This value is determined by dividing the height of the erythrocytes by the total height of the blood in the tube and multiplying by 100. • Hematocrits vary but there is a range of values that is considered normal. Average hematocrit values are: • males.......... 40-50% • females....... 38-45% • athletes........ > 50% • Any activity or condition that consistently lowers oxygen levels in the blood will cause an increase in erythropoesis and a subsequent rise in the hematocrit.

17. How Blood Clots • Blood Clots on the outside of the body to stop the flow of blood through cuts. • They can also form on the inside of the body due to trama or disease. • http://www.biosbcc.net/doohan/sample/htm/Hemostasis.htm

18. Blood Clots Blood clots have a 13 step process to form blood clots. Injuries like stabbings and shootings can cause secondary blood clotting that can kill the individual. Lipid deposits can cause thinning of the blood vessels and beginning points of clots.

19. Blood Clotting from injury

20. More than 15 blood antigen systems have been identified, but the A-B-O and Rh systems are the most important. This system determines which blood can be given to a patient and also helps to identify and eliminate suspects in criminology. • These blood types are due to antigens and antibodies that can cause blood to clot or coagulate into small clumps. • An individual that is type A has A antigens on his/her red blood cells, type B has B antigens, AB has both A and B antigens, and type O has neither A nor B antigens. The antibodies are in the blood and determines which blood can be used in a transfusion attempt. • Rh factor is determined by the presence of another antigen, the D antigen. Blood Typing

21. People having the D antigen are Rh positive; those not having the antigen are Rh negative. • For every antigen there is a specific antibody that will react with it to form clumps known as agglutination. • Thus, if serum containing anti-B is added to red blood cells carrying B antigen, they will immediately react. Blood Typing

22. Blood Agglutination Explained

23. The term serology is used to describe a broad scope of laboratory tests that use specific antigen and serum antibody reactions. • The identity of each of the four A-B-O blood groups can be established by testing the blood with anti-A and anti-B sera. Serology

24. The concept of specific antigen–antibody reactions has been applied to immunoassay techniques for the detection of drugs of abuse in blood and urine. When joined due to incompatibilities they form agglutinations or precipitates Serology Antigen-Antibody Reaction Antigens and Antibodies freely flowing in the blood.

25. A number of immunological assay techniques are commercially available for detecting drugs through antigen-antibody reaction. • One such technique, the enzyme-multiplied immunoassay technique (EMIT), is used by toxicologists because of its speed and high sensitivity for detecting drugs in urine. • In a typical EMIT analysis, antibodies that will bind to a specific drug are added to the subject’s urine. • Other immunoassay procedures are also available, such as radioimmunoassay (RIA), which uses drugs labeled with radioactive tags. Immunoassay

26. When an animal, such as a rabbit or mouse, is injected with an antigen its body will produce a series of different antibodies, all of which are designed to attack some particular site on the antigen of interest. Antigen-Antibody Reaction

27. This collection of antibodies is known as polyclonal antibodies. • Alternately, a more uniform and specific collection of antibodies designed to combine with a single antigen site can be manufactured. • Such antibodies are known as monoclonals. Antigen-Antibody Reaction

28. The criminalist must be prepared to answer the following questions when examining dried blood: • 1. Is it blood? • 2. From what species did the blood originate? • 3. If the blood is of human origin, how closely can it be associated to a particular individual? • The determination of blood is best made by means of a preliminary color test. Forensics of Blood

29. A positive result from the Kastle-Meyer color test is highly indicative of blood. Hemoglobin causes a deep pink color. • Alternatively, the luminol test is used to search out trace amounts of blood located at crime scenes. • Luminol produces light (luminescence) in a darkened area. Testing for Blood Courtesy of C. Fanning ) Courtesy of C. Fanning Courtesy of C. Fanning

30. Microcrystalline tests, such as the TakayamaandTeichmann tests, depend on the addition of specific chemicals to the blood so that characteristic crystals will be formed. Testing for Blood Cont.

31. Once the stain has been characterized as blood, the precipitin test will determine whether the stain is of human or animal origin. Testing for Blood

32. Testing for Blood • The precipitin test uses antisera normally derived from rabbits that have been injected with the blood of a known animal to determine the species origin of a questioned bloodstain. • Once it has been determined that the bloodstain is of human origin, an effort must be made to associate or dissociate the stain with a particular individual. • DNA analysis has allowed forensic scientists to associate blood to a single individual.

33. A-B-O vs. DNA • Prior to the advent of DNA typing, bloodstains were linked to a source by A-B-O typing and the characterization of polymorphic blood enzymes and proteins. • This approach has now been supplanted by the newer DNA technology. • DNA analysis has allowed forensic scientists to associate blood and semen stains to a single individual.

34. Heredity and Paternity • The transmission of hereditary material is accomplished by means of microscopic units called genes, located on chromosomes. • Alternative forms of genes that influence a given characteristic (such as eye color or blood type) are known as alleles. • Paternity testing has historically involved the A-B-O blood typing system, along with blood factors other than A-B-O. • Currently, paternity testing has implemented DNA test procedures that can raise the odds of establishing paternity beyond 99 percent. How its done.

35. Punnett Squares • Punnett Squares are used to show the crossing of the genes from the mother & father to produce the genotype of blood type. • The Phenotype is the physical presentation of the organism after birth.

36. Blood Type Genetics Example

37. Testing for Seminal Stains • Many of the cases sent to a forensic laboratory involve sexual offenses, making it necessary to examine exhibits for the presence of seminal stains. • The best way to locate and at the same time characterize a seminal stain is to perform the acid phosphatase (an enzyme secreted into seminal fluid) color test. • A purple color indicates acid phosphatase enzyme.

38. Testing for Seminal Stains Semen can be unequivocally identified by either the presence of spermatozoa or of p30, a protein unique to seminal plasma. Forensic scientists can successfully link seminal material to an individual by DNA typing.

39. Rape Evidence • The rape victim must undergo a medical examination as soon as possible after the assault. • At that time the appropriate items of physical evidence including clothing, hairs, and vaginal and rectal swabs can be collected for subsequent laboratory examination. • All outer and undergarments should be carefully removed and packaged separately in paper (not plastic) bags. • Bedding, or the object upon which the assault took place, may also be carefully collected.

40. Rape Evidence • If a suspect is apprehended within 24 hours of the assault, it may be possible to detect the victim’s DNA on the male’s underwear or a swab of the suspect. • Items routinely collected from the suspect include all clothing, pubic hair, and a blood sample or buccal mouth swab for DNA typing. • The forceful physical contact between victim and assailant may result in a transfer of such physical evidence of blood, semen, saliva, hairs, and fibers.