Examination of Blood

By : Gan Quan Fu, PT, MSc. Human Anatomy (Batch 3)

Content• Introduction

o Historyo Locard’s Exchange Principleo Blood Evidenceo Forensic Value of Blood

• Nature of Bloodo General Characteristicso Roleso Red Blood Cellso Serum

• Blood Grouping• Individualization of Blood• Blood Analysis

o General Questiono Screening Testo Determination of Sex, Race and Age

• Crime Sceneo Wet VS Dry Bloodo Nature of Crimeo Cause of Deatho Blood Splashes

• Collection of Blood Stain• References

INTRODUCTION

HistoryProf. Dr. Edmond Locard 

o (13 Dec. 1877 – 4 May 1966) o Pioneer in forensic science

who became known as Sherlock Holmes of France.

o Formulated the basic principle of forensic science: "Every contact leaves a trace".

o Became known as Locard's exchange principle

Locard’s Exchange Principle

“ Wherever he steps, whatever he touches, whatever he leaves, even unconsciously, will serve as a silent witness against him. Not only his fingerprints or his footprints, but his hair, the fibers from his clothes, the glass he breaks, the tool mark he leaves, the paint he scratches, the blood or semen he deposits or collects. All of these and more, bear mute witness against him. This is evidence that does not forget. It is not confused by the excitement of the moment. It is not absent because human witnesses are. It is factual evidence. Physical evidence cannot be wrong, it cannot perjure itself, it cannot be wholly absent. Only human failure to find it, study and understand it, can diminish its value”

- Professor Dr. Edmond Locard

Blood Evidence• Most well-known and significant

evidence in modern criminal justice system. Because:o It can link a victim to a suspect

(via Locard’s Exchange Principle).o Bloodstain patterns reveal great

deal about position and movement during crime.

o It managed to destroy self-defense arguments of suspects.

• There's no substitute for it, whether for medical or forensic purposes.

Forensic Value of Blood• Blood has always been

considered class evidence in forensic Science.

• Individualized blood evidence is possible in the near future.

• In some cases, forensic serologists were able to link a single perpetrator to a bloodstain with strong probability estimates.

NATURE of BLOOD

General Characteristics • Slightly alkaline fluid • Circulates throughout our bodies, nourishing our

cells, transporting oxygen and waste. • Fluid portion of blood consists of plasma and serum .• Non-fluid portion consists of red blood cells which

outnumber white cells by five hundred to one• Blood is composed of:

o Watero Cellso Enzymeso Proteinso Other inorganic substances

Roles• Medical scientists are more interested in

white cells.• Forensic scientists are more interested in

red cells and secondly serum.

Red Blood Cells• Most prevalent blood cells in

the human body.• Deliver oxygen from the

lungs to the body’s tissues. • Forensic analyst search for

smaller chemical substances residing on their surfaces (ie. antigens), which also tend to have important forensic implications.

Serum• Characterized by its yellow hue,

and contains white blood cells and platelets.

• Forensic analysts able to determine the freshness of a blood sample by examining serum (it clots several minutes after exposure to air).

• A centrifuge can be used to separate the clotted material from the serum portion.

• Serum contains antibodies , proteins floating in blood fluid, which have significant forensic implications.

BLOOD GROUPING

Blood GroupingA-B-O system of blood typing was discovered in 1901 by Dr. Karl Landsteiner, an Austrian biologist and physician.  (June 14, 1868 – June 26, 1943)

o Developed the modern system of classification of blood groups from his identification of the presence of agglutinins in the blood.

o 1909 Discovered polio virus. o 1930 Received Nobel Prize in

Physiology or Medicine. o 1937 Discovered Rhesus factor.o 1946 awarded a Lasker Award and is

recognised as the father of transfusion medicine.

Blood Grouping• Late 1930s, a series of antigen-

antibody reactions discovered in blood, the most common ones being ABH, MN, Rh, and Gm (gamma glouburin marker).

• There are more than 256 antigens, and 23 blood group systems based on association with these antigens.

• A fundamental principle of serology is that for every antigen, there exists a specific antibody.

• All blood groups are defined by the antigens on their red blood cells and the antibodies in their serum.

• ABO blood group antigens present on red blood cells

• IgM antibodies present in the serum.

BLOOD TYPING

Blood Typing• Blood typing requires 2 antiserums:

1. anti-A 2. anti-B.

• Insert a droplet of these antiserums in samples of blood can determine which samples maintain normal appearance and which become clotted, or agglutinated under microscopic examination.

‘A’ blood agglutinated by anti-A serum. ‘B’ blood agglutinated by anti-B serum.

‘AB’ blood by both.

‘O’ blood by neither.

Rhesus Factor• Can be categorized using Rh

(Rhesus disease) factor. • Positive Rh factor, means the

blood contains a protein that is also found in Rhesus monkeys. 

• Approximately 85% of the population has a positive Rh factor.

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

INDIVIDUALIZATIONof

BLOOD

Individualization of Blood

• Potential for the individualization of blood is based on the typing of proteins andenzymes.

• Blood proteins and enzymes have the quality of being polymorphisms or iso-enzymes, which means they exist in several forms and variants.

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

Secretors• 1925 Blood-related discovery valuable to forensic

science was made. • Approximately 80% of the human population was

found to be "secretors” individuals whose specific types of antigens, proteins, antibodies, and enzymes characteristic of their blood can be found in other bodily fluids and tissues. 

• In the case of a secretor, investigators can conclude the blood type by examining the saliva, teardrops, skin tissue, urine, or semen.

• In a rape case, for example, where the perpetrator is a secretor, potential suspects can be narrowed down through blood type analysis.

BLOOD ANALYSIS

General QuestionsFive specific questions as guidelines for determining the nature of a crime:

1. Is the sample, blood?

2. Is the sample, animal blood?

3. If the sample is animal blood, from which species did it come from?

4. If the sample is human blood, what type is it?

5. Can the sex, age, and race of the source of blood be determined?

Screening Test (Is it Blood?)1. Luminol Test

o To reveal these traces with a light-producing chemical reaction between several chemicals (ie hydrogen peroxide) and hemoglobin, an oxygen-carrying protein in the blood.Benzidine Test

2. Phenolphthalein( Kastle- Meyer’s) Testo Phenolphthalein is colourless when it's reduced (has electrons), and

pink when it's oxidized with hemoglobin (no electrons)

3. O-tolidine Testo a test for the presence of hemoglobin in feces or urine. The O-

toluidine can be in solution, in a tablet or impregnated onto absorbent paper. A positive test is the development of a green to blue color.

4. Leucomalachite Green Test

Luminol Test

Phenolphthalein( Kastle- Meyer’s) Test

Screening Test (Human or Animal

Blood?)

• Precipitin Test• Double gel diffusion technique of

Ouchterlony(anti-human hemoglobin tests)• ABAcard HemaTrace Test

ABAcard HemaTrace• Method of choice today• An immuno chromographic one-step test for the

detection of human blood• HemaTrace uses anti-human hemoglobin (Hb)

antibodies to provide a means of detection for the presence of human blood

• Is coated on test strip and has a detection limit of 0.07 ug Hb

Determination of Sex. Race and Age

• Sex Determinationo Presence of Davidson’s body in leukocyteso Can be identified by as early as 7 weeks into

pregnancy; mother’s blood for Y-chromosome specific.

o DNA (Positive indicates male fetus)

• Raceo Certain racial genetic markers involving protein and

enzyme tests can help establish race.

• Age Determinationo Fetal Hb indicates fetal blood; also alpha- fetoprotein

DNA Test EDTA is the anticoagulant of choice for blood collection for DNA extractions

o It inhibits DNase activity and does not introduce volume changes

o Irrespective of the anticoagulant, the Vacutainer tube should be inverted several times to mix the blood

o Sample should be shipped as early as possible within 24 hrs( delay > 3days degrade DNA )

CRIME SCENE

Wet VS. Dry Blood• Wet blood is more significant than

dried blood because the forensic scientist can perform more tests in order to gain insight to the happenings of the crime. (ie. alcohol and drug content can be determined from wet blood only. )

• Blood begins to dry after 3 to 5 minutes of exposure to air.

• As it dries, it changes colour from deep red to brown and black.

• Blood can be categorized into pools, drops, smears, or crusts.

Nature of Crime• Murder

o Blood on weapon can be matched against the blood of the victim

o Blood stains on the clothing's or person of the accused can be matched with blood of the victim

o Hair roots found on weapon’s can be matched with blood of victim’s and accused

• Accidental deaths( RTA etc)o Blood stains on the tyre, radiator grill, and other part of the

offending vehicle may connect victim and it• Sexual crimes

o Blood stains on thigh and private parts of the victims • Paternity disputes

o HLA, DNA• Blood of pregnancy/abortion

o Presence of chorionic gonadotropin ( early stages)o Heat –stable alkaline phosphatase( late stages)

Cause of Death• Death due to Asphyxia’s :

o Dark fluid blood in heart chambers• Death due to drowning:

o Alteration in Chloride content( Gettler’s test)• Death due to poisoning:

o Carbon monoxide poisoning: cherry pink color of blood

o Cyanide Poisoning: Brick red color of bloodo Heavy metal poisoning (ie. lead): anemia,

basophilic stippled cells

Blood Splashes1930s, Scottish pathologist John Glaister classified blood splashes into 6 distinct types:

1. Drops on a horizontal surface.2. Splashes, from blood flying through

the air and hitting a surface at an angle.

3. Pools around the body, which can show if it's been dragged.

4. Spurts from a major artery or vein. 5. Smears left by movement of a

bleeding person. 6. Trails, either in form of smears when

a bleeding body is dragged, or in droplets when it is carried.

Categories of Blood Stain1. Passive (dripping)2. Transfer (smearing)3. Projected

o Occur in shootings, trauma from blunt weapons, hacking, or slashing attacks.

Passive Spatter Projectile Spatter Contact Spatter

Passive Spatter: Blood that drips because of gravity and as just an after effect of the violence is called the passive spatter. This can range from isolated drops of blood to stagnated pools.

Blood Spatter

Projectile Spatter: Blood that drops as a result of an external impulse is called projectile spatter. This includes blood projecting from various wounds as a direct result of the violence.

Smearing: This includes marks which have been left as imprints of something drenched in blood coming in contact with a target area.

Projected Blood Stain Analysis• 2 Important Determinations:

o Direction of splattero Angle of impact with surface

• sin = (width drop / length drop)

Arterial or Venous Blood• Arterial blood

o Copiouso Bright red in colour o Spurting

• Venous bloodo Dark redo Oozes out gradually

Ante mortem / Post mortem Blood • Ante mortem

o Due to presence of fibrin blood effused during life can be peeled off in scales upon drying

• Post mortemo Blood flowed after death tend to break

into a powder upon drying

Collection of Blood Stain

Precaution• Wear protective clothing, gloves, masks, and/or

eye protection as the situation warrants

• Photographs should be taken from all angles

Liquid / Fresh Stain• A clean white filter paper or a piece of clean

white cloth; a control filter paper should be also sent if object is porous, obtain a portion of the unstained area as well

Old and Dried Stain• Should be carefully scraped if on a immovable

object. • Scrapings placed in clean containers or envelopes

sealed and labelled.• On clothes or fibres.... Scrapped off or a fragment

of the material cut, collected in paper bags, sealed & labelled.

• Dried blood stains on weapons, garments, etc. can be left intact and entire object submitted.

• Tape lifting bloodstains o If finger prints present

Solvent of Blood• 10 % solution of potassium cyanide• 10% solution of glycerin in distilled water (most

common)• A weak solution of ammonia

• Methodso Sample can eluted by rubbing the stained area

with cotton swab moistened with distilled water, the swab is air dried and not heated. Placed in a sterile swab tube, labeled and sealed

o Arrange and write the swabs and collected samples in sequence of collection

Preservation of Blood• Sodium fluoride • Potassium oxalate / lithium or ammonium

oxalate, • EDTA • Citrate( sodium citrate)• Sodium Iodo acetate • 4 degrees Celsius Temperature

Conclusion• Blood is an important evidence in crime scene

and must be carefully detected and collected.• Blood screening is used generally to screen out

the suspects not involve and to provide a more solid evidence to determine the criminals.

References• Bureau of Criminal Apprehension. Available at:

https://dps.mn.gov/divisions/bca/bca-divisions/forensic-science/Pages/forensic-programs-crime-scene-luminol.aspx

• Science Service Justice. http://www.nfstc.org/pdi/Subject02/pdi_s02_m02_02_b.htm

• UCSB Scienceline. Available at: http://scienceline.ucsb.edu/getkey.php?key=2777