CHAPTER III Blood and Blood Stains
CHAPTER III Blood and Blood Stains
CHAPTER III Blood and Blood Stains
The significance of blood and bloodstains as evidence in crimes of violence is very obvious such
that we need not place emphasis on this. The tests for the identification of blood are employed as an
important part of the routine investigation in many case of violent death. The specimen usually submitted
fresh blood or fluid blood, dried blood and clotted blood. Very often it is brought to the laboratory in the
form of dried red or brown stains on weapons, clothing or other objects.
BLOOD – has been called the circulating tissue of the body. It is referred to us highly complex mixtures
of cells, enzymes, proteins and inorganic substances. It is the red fluid of the blood vessels. Blood is
opaque. On treatment with ether, water or other reagents becomes transparent and assumes lake color. It
is faintly alkaline. Normal pH is 7.35 to 7.45.
COMPOSITION OF BLOOD
c. Blood Platelets or THROMBOCYTES – Cells that are produced by the bone marrow and
are necessary for proper clotting of blood. Normally responsible for the retraction of
blood clot.
2. 55 % Plasma – The fluid or liquid portion of blood where the cells are suspended. It is principally
composed of:
a. Water (90 %)
b. solid (10%) – largely protein in nature and consists of albumin, several globulins and
fibrinogen
Albumin- the most abundant protein in the blood. It binds with many drugs.
Globulins- Have an important role in the immune mechanism of the body. The globulins carry drugs as
well as sex and thyroid hormones, lipids and iron.
Fibrinogen- the soluble precursor of fibrin which forms blood clot.
PLASMA – the yellowish fluid of blood in which numerous blood corpuscles are suspended. A straw-
Yellow liquid formed when blood to which an oxalate has been added to prevent clotting is
allowed to stand.
SERUM – A straw-yellow liquid formed when clotted blood is allowed to stand for sometime and the
blood contracts.
4. As evidence in the determination of the direction of escape of the victim or the assailant.
5. As evidence in the determination of the origin of the blood.
6. As evidence in the determination of the approximate time the crime has committed.
1. PRELIMINARY TEST – Determines whether the stain contains blood or another substance. It is
used to demonstrate the presence of blood. It determines whether visible stains do or do not
contain blood.
2. CONFIRMATORY – test that possibly identify blood. Determines whether bloodstain really
contains blood.
3. PRECIPITIN TEST – Determines whether the stain is of human or animal origin. Determines
whether blood is of human or non-human origin, and if non-human, the specific animal family
which it originated.
1. BENZIDINE TEST
2. PHENOLPHTALEIN TESTS also known as KASTLE-MEYER TEST.
3. GUAIACUM TEST also known as VAN DEEN TEST, DAY’s or SCHONBEIN’S TEST
4. LEUCOMALACHITE GREEN TEST
5. LUMINOL TEST
Extremely sensitive tests that can be apply to minute stain. For many years the most commonly
used preliminary test for blood. Its use has generally been discontinued as it is known carcinogen. A very
delicate test and will detect blood when present in dilution of 1:300,000 parts. The benzidine test never
fails to detect blood even when very old, decomposed stain with all sorts of contamination is examined.
This test is more sensitive than Guaiacum test and is valuable as a negative result. If the stain react
negatively it is not blood. The positive result is only indicative that blood may be present.
REAGENT: a. Benzidine solution (small amount of powdered benzidine dissolved in glacial acetic acid)
b. 3 % solution of hydrogen peroxide.
PROCEDURE: Place a small fragment/portion of the stained material on a filter paper. Add a drop of
benzidine solution and then a drop of hydrogen peroxide solution.
PHENOLPHTHALEIN TEST
An alternative test to benzidine test. It can detect blood in a dilution of 1:80,000 parts. A positive
result with this test is highly indicative of blood. The negative is therefore valuable and is conclusive as to
the absence of blood.
REAGENT:
A. Phenolphthalein solution(1 to 2 g. of Phenolphthalein to 100 ml of a 25% potassium hydroxide
in water added with 1 g of Zn powder heated until colorless.)
LIMITATION OF THE TEST: The test is also given by copper salts, potatoes and horseradish.
An important presumptive identification test for blood. The reaction of luminal with blood results
in the production of light rather than color. By spraying luminal reagent unto a suspected item, large area
can be quickly screened for the presence of blood stains. The sprayed object must be located in a
darkened area while being viewed for the emissions of light. Luminol test is extremely sensitive test. It is
capable of detecting bloodstains diluted up to 10,000 times. Luminol is known to destroy many important
blood factors necessary for the forensic characterization of blood, so its use should be limited only to
seeking out blood invisible to the naked eye.
PEROXIDASE – is an enzymes that accelerates the oxidation of several classes of organic compounds by
peroxide.
1. Microscopic test
2. Microchemical test or Microcrysttaline test
3. Spectroscopic test
VISIBLE RESULT:
1. Mammalian red blood cells- circular, biconcave discs with nucleus. Appear as characteristic non-
nucleated disc. Exception is camel and closely related animal as llama whose red blood cells are
oval but also without nucleus.
2. birds, fish, and reptile red blood cells- larger, oval and nucleated
3. amphibians red blood cells – are larger than mammals, oval and nucleated
4. lamprey eel red blood cells – circular and nucleated
The test depends on the addition of specific chemicals to the blood so that characteristic crystals
with hemoglobin will be formed.
A delicate test for the presence of hemoglobin. The test depends on the addition of specific
chemicals to the blood so that characteristic crystals of hemoglobin derivatives will be formed.
The most delicate and reliable test for the determination of the presence of blood in both old and
recent stains. This test is performed by means of an optical instrument known as SPECTROSCOPE, an
optical instrument for forming and examining spectra.
PROCEDURE: Dissolve bloodstain in water or saline solution. Place in a small chamber (glass) with
parallel sides so arranged that the rays of light will pass directly through it. The chamber is placed in the
spectroscope and the instrument is so adjusted that the spectrum is clearly visible.
POSITIVE RESULT: Upon absorbing some of the rays from the spectrum, it produced characteristic dark
colored bands which vary with the type of blood pigment. Example: oxyhemoglobin is marked by two
bands, hemoglobin-broad band; carboxyhemoglobin its spectrum similar tooxyhemoglobin.
The precipitin test is the standard test used to determine whether the stain/blood is of human or
animal origin. The precipitin test is very sensitive and requires only a small amount of blood for testing.
Human bloodstain dried for as long as 10 to 15 years and longer may still give a positive precipitin
reaction. Even extracts of tissues from mummies four to five hundred years old have given positive
reaction with the test. Experience has shown that human bloodstains diluted by washing in water and left
with only a faint color may still yield a positive precipitin reaction.
REAGENT: Precipitin/antiserum
PROCEDURE: Scrape off bloodstain if on hard material. Powder the scraping and extract with saline
solution. If the stain is on cloth, paper or similar material, cut a small portion and then place in a test tube
and add extract with saline solution. Allow mixture to stand overnight. Centrifuge to clean the solution.
Dilute with saline solution. Layer an extract of the bloodstain on top of the human antiserum/precipitin in
a capillary tube.
POSITIVE RESULT:
1. Development of a white cloudy line at the contact point of the fluids that appears
immediately or within one or two minutes.
2. Human blood, or for that matter, any protein of human origin in the exact will react
specifically with anti-bodies present in the serum as shown by the formation of cloudy
ring or band at the interface of the two liquids.
If the specimen is human blood the next question is did it come from the victim, the accused or
from other persons? So the origin of blood or bloodstains will be determined by the identification of the
blood groups to which it belongs, in short to what blood group does it belong? This identification is
carried out on both fresh blood and bloodstains. Human blood of all races can be divided into definite
groups.
In the blood grouping of fresh blood A-B-O System is used. It was Landsteiner who discovered
the four blood groups namely Group O, Group A, Group B, and Group AB. He named the four groups on
the basis of the agglutinogen or antigen content of the red blood cells. Antigens are characteristic
chemical structures or “principles” that are found on the surface of each red blood cell which stimulates
the production of agglutinins. There are two different agglutinogens classified as agglutinogen A and
agglutinogen B. On the other hand serum contains proteins or “principles” known as antibodies or
agglutinins which cause agglutination or clumping together of the red blood cells. They are antitoxin
substance within the body which reacts when confronted with a specific antigen to protect the system.
There are two different agglutinins classified as anti-A and anti-B in the serum. Agglutinogen A and B are
present at birth while agglutinins are demonstrable in about 50% of newly born infants. If an individual
belongs to group A this indicates that his red blood cells has antigen A located on its surface. Similarly all
group B persons have antigen B, all group AB persons have antigen A and Antigen B and all group O
persons have neither antigen A nor antigen B.
When the serum of group A blood was examined, anti-B was found present and no anti-A. Similarly
Group B blood contains only anti-A, Group O has both anti-A and Anti-B and Group AB blood contains
neither Anti-A nor Anti B.
Knowledge of the laws of genetics will make it easier to understand the principle involved in the
inheritance of blood groups. The inheritance of human blood groups is predetermined by the presence or
absence in the chromosomes of two factors or genes called gene A and gene B. Since each body cell has a
pair of chromosomes, each of which carries or fails to carry one of these factors, an individual’s genetic
constitution may be represented by AB, AA, AO, BB, BO, or OO where O represents the absence in the
chromosomes of either the A or B factor. In the joining of the ovum and spermatozoon during
fertilization, a new pair of genes is formed corresponding to the gene found in the chromosomes of the
parents called zygote. If the genes are homozygous or pure i.e. they are alike and they are the same in
both the father and mother, the characters are transmitted unchanged from generation to generation. If the
two genes are not the same which is called heterozygous or hybrid a new combination will arise in the
next generation.
Beinstein’s theory postulates the presence of three allelic genes A, B, and O. According to him
the blood group of any individual is determined by combination of A, B, and O in a particular pair of
chromosomes. One gene is derived from the father and the other gene from the mother. Genes A and B
are dominant over gene O. Gene A and B determine the presence of the corresponding agglutinogens,
while O determines their absence. The possible combination of these three genes arranged in pairs gives
rise to six different genotypes corresponding to the four phenotypes or the blood groups. There are ten
different matings possible between the four blood groups.
DEFINITION OF TERMS
1. GENE- any of the complex chemical units in chromosomes by which hereditary characters are
transmitted. Occurs in pair. A factor occurring singly in a gamete. There are two genes or factor
called gene A and gene B. These are found in the chromosomes. Since chromosomes go in pair,
each of which carries or fails to carry one of these genes and an individual’s genetic constitution
may be reprersented by AA, AB, BB, BO, AO, OO which are called genotypes, where O
represents the absence in the chromosomes of either the A or B gene. Gene is responsible for the
transmission of hereditary characteristics.
2. CHROMOSOMES- any of the microscopic rod-shaped bodies bearing genes responsible for the
transmission of hereditary characteristics. They are observed to occur in pairs.
3. PHENOTYPES- term used to denote the expression of the inherited characteristics as found in
the individual. Actually the blood groups.
4. GENOTYPES- are paired genes. It is either homozygous or heterozygous.
5. HOMOZYGOUS GENOTYPE OR PURE GENOTYPE- paired genes are similar
6. HETEROZYGOUS GENOTYPE OR HYBRID- paired genes are dissimilar or not alike.
7. GAMETE- sexual cells; reproductive cells that unites with one another to form cell that develops
into a new individual.
8. SPERM CELL OR MICROGAMETE- male sexual cell
9. EGG CELL OR MACROGAMETE- female sexual cell.
10. ZYGOTE- pair of genes occurring in a gamete produced during fertilization. Cell formed by the
union of an ovum and a sperm.
11. ALLELES- pairs of contrasting genes which determines the expression of the inherited
characteristics of an individual.
THE M-N SYSTEM OF BLOOD GROUP
In 1927 Landsteiner and Levine discovered two new agglutinogens in human red blood cells
which define three types of blood, namely type M, type N, type MN. These are independent of the
agglutinogens A and B. The human sera however do not contain natural agglutinins for N, and rarely
agglutinins for M. The agglutinins can be demonstrated only by heter-agglutination reaction with
appropriate immune rabbit sera. So anti-M and anti-N were produced by heter-agglutination reaction and
just like Anti-A and Anti-B it determines the presence of agglutinogen M and N in the red blood cells.
Six different mating are possible between the three blood types. Types MN is always heterozygous.
The heredity of agglutinogens M and N according to Landsteiner and Levine depends upon a single pair
of allelic genes M and N which give rise to the three genotypes MN, MM, and NN corresponding to the
three phenotypes MN, M, and N respectively.
Questions of illegitimacy and relationships in many cases may be solved by means of the blood
groups as determined by the agglutinogens A, B, M, and N.
1. Determination of whether accused of fathering a child out of wedlock could or could not be its parents.
2. Determination of whether a child born of a married woman could or could not have been fathered by
her legal spouse.
3. Determination of whether a child could or could not belong to a given set of parents in the case of
accidental interchange of infants in hospital.
4. Determination of whether a child who has been lost and later recovered after a long interval could or
could not belong to a given set of parents.