Advanced forensic biology

Unit – 1
Forensic biology :
• It is the scientific analysis of biological evidence to provide objective
information on legal matters or those that pertain to criminal and civil law.
• Forensic Biology is a sub-discipline of forensic science. It applies the
knowledge of biology to identify and analyze the biological evidence obtained
from the scene of a crime or from victim or suspect(s) to establish the fact
that a crime has taken place.
• It is a broad discipline that includes various areas of specialization such as
DNA analysis, forensic anthropology, forensic pathology, forensic
entomology, forensic odontology, forensic botany, forensic serology, and
forensic microbiology.
• Biological evidence such as bodily fluids or tissues that may be found at
crime scenes can be analyzed through serological and DNA typing. Typing
requires detection and screening of the biological evidence such as blood,
semen or saliva, extracting the DNA from a specimen, amplifying specific
regions of the DNA using the polymerase chain reaction (PCR), and typing
the resulting PCR products to determine a DNA profile.
• The DNA profile from the evidence is then compared to known profiles from
suspects, victims or data base samples to determine the significance of the
result. Samples containing mixtures will require additional interpretation to
infer individual donor allele designations.
• Forensic biologists must also assess the statistical significance of their
results, write reports and testify in court.

A Brief History of Forensic Biology

• Karl Landsteiner: In 1901, he discovered that the blood could be grouped into
different categories, A, B, AB, and O.
• Dr Leone Lattes: In 1915, he discovered a simple procedure to determine the
blood group of the dried bloodstains. This technique was adopted for
criminal investigation.
• Sir Alec Jeffreys: In 1984, he developed the DNA fingerprinting technique to
examine the variations in the genetic code, which, can be used to distinguish
one individual from another.
• Kary B. Mullis: In 1993, he developed the PCR technique that was used to
amplify the samples of DNA fragments. This technique was useful for
amplifying the DNA sample obtained from the scene of a crime in an
extremely minute amount, degraded state, and a mixture of body fluids of
two or more people.
Development :
• The establishment of a United States national DNA database, the Combined
DNA Index System or CODIS, has facilitated the ability to compare DNA
profiles from unknown biological crime scene evidence to DNA databases of
known convicted criminals leading to “cold hits” or to DNA left at other
crime scenes resulting in the ability to link cases. Many other countries
have DNA databases with some of the same genetic markers permitting
international database searches.
• By comparison of the DNA profile from crime scene samples to known
samples, the results can serve to link victims and suspects with the crime
scene, or can exclude a suspect from association with that crime.
• Additionally, scientific analysis of biological evidence may provide unbiased
information to substantiate case circumstances, corroborate or refute an
alibi, and/or identify a weapon used in a crime.
• Cases may include non-human samples such as botanical, fungal,
entomological or zoological specimens that can also be used to link victims
and suspects with each other or to the crime scene.
Significance And Application Of Forensic Biology
The field of forensic biology deals with the examination of the evidence
pertaining to living beings, and their associated biological materials, commonly
found at the scene of a crime.
This deals with the following works:
• Examination, identification, of biological fluids like blood, urine, semen,
sweat, saliva, milk, teas, juice, and determining their origin either come
from the human, plant or animal sources.
• Examination and identification of materials like wood, hair, fibre, fecal
matter, nails, bones, teeth, leaves, seeds, pollen, and other plant
materials.
• Examination and detection of microbial organisms in food, animal, human,
and water samples.
• Examination and identification of diatoms in the water, human viscera,
and food items.
• Determination of the group of blood, semen, sweat, saliva if found to be of
human origin.
• Determination of paternity and maternity of an individual by DNA profiling.
• Determining the time since death by examining the bones, and teeth.
• Determining the age and sex of human beings by examining the skeletal
remains.
• Determining the time since death, and manner of death by examining the
insects found on and near the deceased.
Sub-Fields Of Forensic Biology
• Forensic DNA Analysis - DNA or deoxyribonucleic acid is the most relevant
evidence which can be obtained from the scene of a crime. The DNA analysis
has numerous applications like identification of unknown biological remains
 (like blood, seminal fluid, sweat, bones, hair with roots, tooth pulp, and
others) and paternity testing, missing person identification, cases of
impersonation, etc.
• Forensic Serology- Forensic serology the scientific study or diagnostic
examination of blood serum, especially with regard to the response of the
immune system to pathogens or introduced substances) deals with the
preliminary and confirmative analysis of biological fluids. The knowledge of
serology will help in determining the origin of the biological fluids, whether
of human, vegetable, or animal origin. It also helps in determining the blood
type.
• Forensic Botany- It involves the identification of the source of plant matter
like leaves, stems, pollen grains, seeds, and fibres found around or over the
deceased, which can help in determining the place of commission of a
crime.
• Forensic Anthropology- Forensic Anthropology is the sub-field of forensic
biology which deals with the collection and identification of skeletal remains
either they are from animal or human origin. The field of anthropology is
useful in a criminal investigation as the knowledge of anthropology helps in
the identification of a deceased by helping in the determination of race, sex,
age from skeletal remains.
• Forensic Odontology- Tooth enamel is the hardest part of the human body,
which takes many years to degrade and decompose when the body is
buried. The teeth obtain at the scene of a crime or from the buried remains
can be used to link the culprit or victim. The examination of bite marks over
the surface of the object, body, vegetable stuff can be useful for the
identification of the criminal.
• Forensic Pathology- It deals with the detailed external and internal
examination of the deceased body through autopsy. This procedure helps in
determining the time since death, manner of death, and cause of death.
• Forensic Entomology- It involves the study of the life cycle of the insects
found near, and inside the decomposed cadaver (dead body) of humans or
animals. This process helps in the identification of the cause of death, time
since death, and place of death. It is associated with the death investigation,
and if the skeleton is obtained, the insect analysis can reveal that the body
was intoxicated with drug or poison.
• Forensic Microbiology- It involves the application of knowledge and
expertise of microbiology for the identification of crime and using microbial
evidence for the conviction of the criminal. The microbial evidence can be
found in the cases of drowning deaths, hospital and clinical acquired
infections, The microbial study can be useful for the identification of a
person associated with biological material such as hair, skin, blood, saliva,
urine, and fecal matter. The microbial study can also be useful for the
determination of the post-mortem interval.
Role Of Forensic Biologist
• The forensic biologist examines blood, and other bodily fluids, bones, hair,
insects, plants, animal remains, diatoms, found at the scene of a crime. The
evidence is analyzed, and the forensic biologist establishes their relation
with the crime in their expert testimony.
• A forensic biologist collects the evidence from the scene of a crime,
analyzes the biological evidence found in close proximity with food, clothes,
soil, water, and other materials.
• The forensic biologist keeps a log of details of the evidence like date, venue,
and time of collection of the evidence and name of the experts to which the
evidence was handled as attention is required because a single mistake can
disturb the integrity, and acceptability in the court of law.
Biological evidence The term biological is derived from the word biology,
which refers to living organisms, whereas evidence is defined as something
legally submitted in a court of law to ascertain the truth of any alleged matter
of fact under investigation. Therefore, biological evidence can be referred as
biological materials or substances such as hair, tissue, bones, teeth, blood,
semen or other bodily fluids.
Types of Biological Evidences-
• Biological evidence comes from various sources of origin. It can be from
human, plants or animals origin. Pollen, algae, fungi and diatom are
examples of evidence from plants. Likewise microbes and insects are
evidence from animals. Biological evidence from humans include blood and
blood stains, semen and seminal stains, saliva, urine, tissues and cells,
bones and organs, hair, teeth, fecal and fecal stains, vomit, stomach
contents, sweat, ear wax etc. Importance of Biological Evidences-
• Help in relating corpus deliciti ( body of offence which shows that a crime
has occurred)
• Helps in victim-to-perpetrator linkage.
• Helps in victim-to-crime scene linkage.
• Perpetrator-to-crime scene linkage.
• Helps in identifying modus operandi of serial offenders.
Blood
Blood is a fluid connective tissue that consists of plasma, blood cells and
platelets. It circulates throughout our body delivering oxygen and nutrients to
various cells and tissues. It makes up 8% of our body weight. Males have 5-6
litres and females 4-5 litres of blood in their body approximately. Blood is
slightly basic, with the pH in the range of 7.35
Blood is very complicated liquid tissue. It is used as transporting medium for
all the substances in the organism. It is to some extent alkaline fluid that flows
in our bodies.
Nature of blood :
The recentness of a blood sample can be determined by inspecting the serum
since it clots quite a few minutes later on exposure to air.
A centrifuge can be used to separate the clotted material from the serum
portion.
In addition, serum contains antibodies, proteins floating in blood fluid, which
have significant forensic effects
Composition of blood

Plasma - The liquid state of blood can be contributed to plasma as it makes up

55% of blood. It is pale yellow in colour and when separated, it consists of
salts, nutrients, water and enzymes. Blood plasma also contains important
proteins and other components necessary for overall health.
Formed elements - Blood consists of cells known as formed elements of
blood. These cells have their own functions and roles to play in the body. The
blood cells which circulate all around the body are as follows:
1. Red Blood cells (Erythrocytes) - The life span of RBCs in humans is
approximately 3 months. Additionally, mature human erythrocytes do not have
nuclei, and therefore lack nuclear DNA. Erythrocytes consist of haemoglobins
which are proteins responsible for the transportation of oxygen.
2. White Blood cells (Leucocytes)- White blood cells are involved in defending
the body against infection. They have nuclei and, thus represent main sources
for nuclear DNA from the blood. WBCs are further divided into granulocytes
and agranulocytes.
Granulocytes contain granules in the cytoplasm. contain enzymes that have
invaded
3. Platelets- These cells are also known as thrombocytes and they play a role
in blood clotting. They aggregate at sites of vascular and blood vessel injury.
Like red blood cells, they lack nuclei.
Importance of blood :
Wet blood is more vital than dried blood as the forensic scientist can
accomplish more test to increase insight to the happening of the crime.
Alcohol and drug content can resolute from wet blood only. Blood starts to dry
subsequently, after three to five minutes of exposure to air and alterations of
color from dark red near brown and black can also be seen. Blood can
characterized into pools, drops, smears or crusts.
Characterization of blood evidence
Blood is the most commonly found evidence at any crime scene so
characterisation of blood evidence is a very crucial step in crime investigation
process.
Characterization of blood
➢ CLASS CHARACTERSTICS associated with a group as in blood group, Rh
factor etc.
➢ INDIVIDUAL CHARACTERSTICS associated with a common source with
certainty as in DNA analysis.
Collection of blood evidence
• Fresh or wet blood should be collected on clean, sterile gauze or cotton
swab and allowed to dry at room temperature.
• Four sampling methods for dried blood -
1. Cutting- For stains on objects that are difficult to submit to the lab. The
cut portion should include unstained portion around the bloodstain.
2. Swabbing- Stain is transferred to a swab which has been moistened
with sterile water or saline.
 3. Scraping- A sharp instrument is used to scrape the stain off a surface
and placed onto a clean paper.
4. Elution- using a small amount of saline or distilled water to dissolve the
dried stain.
Preservation of blood evidence :
➢ The most important consideration for preserving blood evidence from crime
scene is to thoroughly dry the item before packaging and then store in a
cool dry environment.
➢ Wet blood evidence is drawn into a vacutainer tube containing an
anticoagulant such as EDTA.
➢ Dried bood evidence must be packaged in paper containers that are
breathable.
Forensic identification of blood :
➢ Forensic identification of blood includes presumptive and confirmatory
tests.
1. Presumptive tests are used to screen evidence for the possible presence
of blood.
2. Confirmatory tests are the tests which are entirely specific for the
presence of blood.
Scope
blood as evidence holds significance in the criminal justice as it can link a
crime with criminal or exclude on individuals involvement in a crime.
forensic scientist often use techniques to identify blood types (blood typing)
because an individuals blood type isn’t affected by diseases, drugs, climate,
occupation, living conditions, or any other physical circumstances.
Additionally scientist often use techniques to identify blood typing to determine
paternity.
Moreover the pattern of blood stains can help in reconstruction of a crime
scene by conveying information about the relative position and movement of
the criminal and victim at the crime scene
At the crime scene, it is important to establish the type, origin, and other
characteristics of the blood/blood stain.
Preliminary investigation in this regard are to confirm whether the recovered
fluid is blood and whether its origin in human.
The present scenario, DNA technology has replaced the tests for specific
enzymes and proteins from the blood sample.
The DNA typing method is widely accepted in the various courts of law
worldwide. The evidence from blood alone or along with other trace evidences
at the crime scene can play a vital role in solving criminal cases.
Presumptive tests for blood :
➢ The chemistry employed in these assays is an oxidation–reduction reaction
catalysed by heme, a component of haemoglobin. The heme catalyzes
various colourless substrates to undergo an oxidation reaction that results
in a change of color, chemiluminescence, or fluorescence.
➢ In the presumptive assays, hydrogen peroxide is usually employed as an
oxidant. Heme serves as the catalyst for the oxidation– reduction reaction.
In the presence of heme, a colourless substrate is oxidized to a product
with color, chemiluminescence, or fluorescence.
Semen
Semen is found in liquid form, smears or stains or it may be found in vagina,
anus or rectum. Fresh semen is a gel like fluid, which liquefies on exposure to
atmosphere. A normal ejaculation is about 3.5ml, containing about thirty million
sperms. The dry weight is about seven per cent of the liquid weight. The sperm
has definite morphological structure. Its identification in a stain establishes the
presence of semen. The shape and size of human spermatozoon is
characteristic. But the morphology alone does not permit individualization.
Semen of a person does not contain any spermatozoon then it is called as a
spermic semen.
This may be due to some disease or it may be due to vasectomy operation. In
such cases this criterion for the identification of semen is lost. Immunological
test using anti semen sear against seminal plasma are increasingly accepted
as reliable test for aspermic semen. Electrophoresis is becoming popular for
identification of semen.
Composition of semen :
It is a complex mixture of organic and inorganic compounds. Important
constituents of semen from the identification point of view are proteins
including enzymes, blood group factors, choline, fructose, citric acid, uric acid
and zinc. The composition varies from individual to individual. Enzyme, acid
phosphatase found in the semen is in concentration which are significantly
higher than those found in other body fluids. Acid phosphatase offers a very
delicate test for the identification and location of semen stains though positive
identification of semen are not based upon the acid phosphatase test alone.
Choline in semen is used to get crystal tests. Fructose, citric acid and zinc are
more or less absent in other body fluids and hence their detection in semen
should permit its identification but these substances have not been utilized to
any appreciable extent so far.
Locating Semen Semen stains can be located using following techniques:
Visual examination The suspected place or article is examined visually. A semen
stain, if fresh is colourless or pale yellow. It gives its characteristic smell. A
freshly dried stain on wetting also gives the characteristic odour. The stain
becomes stiff and rough to feel on drying. The stiffness does not wear off
unless it is rubbed.
Ultraviolet rays The surface or the article suspected to bear seminal stains
when examined under ultraviolet rays in darkness usually gives fluorescence.
Some other substances also give fluorescence while some other mask it.
Therefore the technique though useful in most of the cases is not infallible. The
fluorescence of semen depends upon the quantity and freshness of the semen.
Therefore, even when a stain shows weak fluorescence, it should be further
explored.
Spectro microscopy examination of the suspected stain may indicate the
characteristic contours and layers like formations in a thick crust of semen
stain.
Phosphatase method The presence of acid phosphatase in semen helps to
search large area and garments for semen stains. The method is: Large filter
paper sheets are dipped in saline. Excess of the solution is removed and the
garments to be examined is pressed against the paper or vice versa. The
outline of the piece of cloth is marked on the paper to facilitate location of the
semen. The filter paper sheet is then sprayed with acid phosphatase reagent.
Color spots indicate the location and size of the seminal stains.
Examination of Semen Stains
Systematic study of seminal stains involves following steps:
• Smell: The smell of fresh or wet seminal stains is characteristic.
• Peel: Dry stains have a rough feel like dried starch solution and have uneven
surface. The contours of the stains are characteristic.
• Fluorescence: semen stains give strong characteristic fluorescence.
Handling, substrata, the quantity and the age of the stain affect it.

❖ Chemical tests
The following chemical test are performed for detection of semen:
• Barberio’s test: A small amount of semen is taken and treated with a
saturated aqueous solution of picric acid. Spermine picrate crystals with
characteristic structure, separate out.
• Florence test: A small amount of semen is treated with iodine in potassium
iodide solution. Characteristic crystals of choline iodide are formed.
• Acid phosphatase test: The acid phosphatase enzyme present in high
concentration in semen liberates phosphate from various compounds
attached to them chemically. This property is utilized in liberating Naphthol
from calcium naphthyl phosphate. Naphthol reacts with a diazonium
compound to give coloured products. Anthraquinon-1-diazonium chloride is
often used in the test. The mixture of various reagents is taken and sprayed
over a wet filter paper, previously pressed against the suspected place
bearing semen stains. Development of red purple color indicates semen, the
technique is extremely sensitive and several years old stains can thus be
detected. This method is however not specific. Recently modifications had
been made which claim to make it more specific.
Morphology of spermatozoon:
The identification of sperms in a stain is considered positive proof of the
seminal nature of the stain. Sperm is a unicellular organism with an oval head
attached to a comparatively long tail. If head and tail of a spermatozoon are
found intact it is not difficult to identify the stain. Intact sperm is often not
available in a dried stain. The sperms may be identified with or without staining.
But staining helps:
• Ordinarily double staining with haematoxylin and eosin is done. In this way
the nose is stained in pink and the rest of the head is stained purple. This
differential staining distinguishes the sperm from any other organism. Head
alone can identify the stain.
• Epithelial cells and leucocytes in excess mask the spermatozoon the
difficulty is overcome by staining the spermatozoon with malachite green
and eosin. The interference is considerably reduced.
Biological tests
Precipitin reaction with anti-human semen serum and specific blood group anti
sera are employed. The former determines whether it is human semen and the
latter determines the blood group of the secretor. The techniques employed in
are similar to those applied for blood grouping
• Individualisation: Semen sample of an individual is unique. With the classical
method it was not possible to establish the individuality but DNA profiling
has made it possible to pin point the source of origin even from the tiniest
speck or even when it is contaminated with vaginal fluid or mixed with
semen from other sources.
• Age of seminal stains: It is difficult to determine the age of seminal stains as
the changes with age are largely determined by the environmental
conditions. The semen is a gel like mass after ejaculation. It liquefies on
standing. On drying, it slowly changes to opaque white mass with rough feel.
After two or three weeks the color changes to pale yellow and then to
brown. Sperms in fresh semen are complete with head and tail. On drying
and with passage of time it begins to disintegrate, the tail separates out
from the head. The head is also affected. The sharp division in differential
staining slowly appears.
• Survival of sperms: The sperms remain motile only for a few hours under
normal conditions. The motility seldom continues after twenty four hours.
But if the semen is frozen, the sperms may remain alive for long periods. It
shows motility when the sample is brought to normal temperature. In humid
and warm climate, the sperms get destroyed in a short while. In dry and
cool climate the dead sperms may be identified even after months. In
vaginal swabs of a living person, sperms may be detected upto about five to
 ten days, though the number of sperms detected goes on decreasing with
passage of time. In dead persons, detection of sperms depends upon the
condition under which the dead body remained. It may be detected in
unpetrified bodies even after a few months. But if the body has putrefied the
sperms are also destroyed.
Semen As A Forensic Evidence
• Semen is a thick, yellowish white, glairy, opalescent, secretion having a
characteristic odour known as seminal odor.It is also called seminal fluid,
fluid that is ejaculated from the male reproductive tract and that contains
sperm cells, which are capable of fertilizing the female eggs.
• Semen also contains other liquids, known as seminal plasma, which help to
keep the sperm cells viable. The total volume of semen for each ejaculation
of a human male averages between 2 and 5 ml. Each ejaculation contains
normally 200 to 300 million sperm. The pH of semen(also referred to as
seminal fluid) is slightly alkaline, ranging between pH 7.2 and 7.4.

As with other forensic exhibits, the investigation of semen is also carried out in
a specific way using various tests.

• First, the screening tests are performed in order to identify whether the
questioned stain is semen or not. Once the preliminary screening tests are
positive, a more detailed confirmatory analysis is carried out to establish
that the stain is indeed semen.

Further exhaustive analysis of seminal stains is performed in order to

individualize the stain to a particular individual.
The identification of one or more sperms is a conclusive proof of the presence
of semen.However, there are difficulties in the identification of sperms due to
several reasons.
1. If a criminal is oligospermic, then his ejaculate may contain only a few
sperms.

2. Another reason could be the condition of Aspermia where no sperms are

produced by the seminiferous tubules of the testes.

3. If the criminal has undergone Vasectomy which is the surgical procedure

for male sterilization to prevent sperm from entering into the seminal
stream.

4. Use of a contraceptives e.g. Condoms

5. In addition to these, there may be other reasons such as sperms trapped in

the clothing not extracted into the testing extracts.

6. Disintegration of sperms can also take place during handling of the

material.

Finding sperm cells in body cavities of a victim of sexual crime is the utmost
priority for obvious reasons. Several research findings indicate that motile and
non-motile sperms can be obtained at different duration in different body
cavities.
COLLECTION & PRESERVATION OF SEMEN

Collection of semen stain has much precaution. Semen converts in to brittle

after drying. So if cloths or other articles are not handled properly or folded it
then spermatozoa are breakdown in to pieces.

• Preservation of semen stain always performed after complete dryness

of stain. If stain is wet then bacterial growth is started and putrefaction
may occur due to presence of protein in semen.
• Any infection in testis due to presence of some bacteria also secretes
liquid like seminal stain from testis which creates doubt about semen.
• It is always preserved in air bag not in airtight bag or plastic bag.
• If stain found on immovable or small article, then preserved whole
article or cloth and sent sample to FSL.
• If stain present on immovable article then sharp knife or scalpel is used
to scratch the stain and packed in clean glass bottle.
• Seminal stain may be present on pubic hair so it is also preserved in bag
or clean plastic tube.
• If seminal stain present on any body part then take cotton swab is taken,
dipped in saline water. Then cotton applies on body and prepare cotton
swab. After that drying the cotton swab and preserved in bag.
• Never touch sample with bare hand because it can contaminate and
spoil the specimen.
• In fresh seminal stain, live spermatozoa are present so keep safe the
cloth and send properly to FSL.
Vaginal fluids :
Vaginal discharge is fluid secreted from tiny glands in the vagina and cervix.
This fluid leaks from the vagina each day to remove old cells and debris,
keeping the vagina and reproductive tract clean and healthy. The amount of
vaginal discharge can vary significantly from person to person.
Composition of vaginal fluid :
Vaginal secretions are continuously released and contain water, nutrients,
electrolytes, and proteins (secretory immunoglobulin A) and are altered by
hormonal and dietary factors.  The normal vaginal secretions are a
heterogeneous suspension of vaginal epithelial cells and lactobacilli in fluid that
emanates from the cervix and vaginal walls. The secretions have a pH of 3.5 to
4.6 and are odourless.
Vaginal cells contain glycogen and are continually shed into the lumen of the
vagina. As the cells autolyze, glycogen depolymerises to glucose, which serves
as an energy source for bacteria known as lactobacilli. Lactobacilli metabolize
glucose to lactic acid, which results in a normal vaginal pH of about 4.0.
Scope :
criminal investigations there are some cases in which identifying the
presence of vaginal secretions provides crucial evidence in proving
sexual assault. However, there are no methods for definitively
identifying vaginal secretions. In the present study, we focused
on Lactobacillus levels in vaginal secretions and developed a novel
identification method for vaginal secretions by relative quantification
based on real time PCR.

FORENSIC ANALYSIS OF VAGINAL FLUIDS :

1. Visual Examination- locates the stains using Alternate light sources.
2. Microscopic Examination- Samples should be taken from the lateral vaginal
walls in the upper third of the vagina to avoid contamination of the sample with
discharge from the cervix. A wet mount (i.e., vaginal secretions diluted with
normal saline) and 10% potassium hydroxide (KOH) preparation should be
performed. The pH should be checked using pH paper that is sensitive in the 3.5
to 6.0 range. During microscopy, the provider should comment on the
presence and nature of epithelial cells, the presence/absence of lactobacilli,
and any potential abnormalities, such as white blood cells, clue cells, hyphae or
budding yeast, or trichomonads.
3. Glycogenotic nucleated squamous epithelial cells (cell type found in vaginal
secretion) is detected by Lugol’s iodine assay or periodic acidSchiff reagent. If
vaginal cells are present, bright magenta colour will be seen.
4. Vaginal acid phosphatase can be differentiated from seminal acid
phosphatase through electrophoresis.

Saliva :
➢ Saliva is a complex biological fluid secreted by acinar cells of the major
and minor salivary glands.
➢ The identification of saliva may be important in the investigation of
criminal and civil cases. For example, saliva evidence can be important in
cases in which oral copulation may have occurred.
 ➢ Additionally, the cells in saliva contain DNA and are often used for forensic
DNA testing.
➢ It is an important discriminating element in forensic biology, acting as an
indicator of salivary gland conditions and toxicological and drug
monitoring.

Biological properties of saliva :

➢ Saliva is a thick, colorless, opalescent fluid that is constantly present in
the mouth of humans and other vertrebrates.
➢ The human salivary glands produce 1.0 to 1.5 litres of saliva daily.
➢ Saliva has a pH normal range of 6.2-7.6 with 6.7 being the average pH. In
the oral cavity, the pH is maintained near neutrality (6.7-7.3) by saliva.
➢ Saliva is an extracellular fluid produced and secreted by salivary glands in
the mouth. In humans, saliva is 99.5% water plus electrolytes, mucus,
white blood cells, epithelial cells (from which DNA can be extracted),
enzymes (such as amylase and lipase), antimicrobial agents such as
secretory IgA, and lysozymes.
Salivary glands and their contribution to saliva volume :
Location of saliva stains :
➢ Saliva is found on clothes, cigarette butt, bottles, cup, handkerchief, etc. In
cases of biting during struggle, assault, kidnapping, hanging, spitting or
tobacco spitting at scene of crime, saliva provides lot of evidentiary data.
➢ Saliva plays an important role in crime investigation because DNA profiling
or grouping test can be performed by saliva due to presence of antigen in
it.
➢ Blood group can detect by saliva although saliva is contaminated by lipstick
or blood.
Collection and preservation of saliva samples :
➢ Fresh or wet saliva samples should be collected on clean, sterile gauze or
cotton swab and allowed to dry at room temperature. SALIVARY GLANDS
SUBMANDIBUAR GLANDS (70%) PAROTID GLANDS (25%) SUBMANDIBULAR
GLANDS (5%)
➢ If chewing gum, tobacco spit, eatables etc. are present at crime scene
then preserve in paper bag as such.
➢ Four sampling methods for dried stains –
1. Cutting- For stains on objects that are difficult to submit to the lab. The
cut portion should include unstained portion around the saliva stain.
2. Swabbing- Stain is transferred to a swab which has been moistened with
sterile water or saline.
3. Scraping- A sharp instrument is used to scrape the stain off a surface
and placed onto a clean paper.
4. Elution- using a small amount of saline or distilled water to dissolve the
dried stain.
Forensic Analysis Of Saliva :
Once the sample is collected and packed, it is immediately sent to the
laboratory for the analysis. There are various tests for the examination of
the saliva.
1. VISUAL EXAMINATION- Visual examination methods such as the use of
alternate light sources can be helpful in searching for saliva stains to
facilitate further analysis. The lighting techniques used to search for
semen stains can be employed in searching for saliva stains. However,
the fluorescence of a saliva stain is usually less intense than that of a
seminal stain.
2. DETERMINATION OF AMYLASE ACTIVITY- Starch iodine test is done.
Iodine is used to test for the presence of starch. The amylose in starch
reacts strongly with iodine to form a dark blue complex, while
amylopectin develops reddish-purple colors. In the presence of amylases,
starch is broken down to mono or disaccharides. Consequently, such
colours do not develop when iodine is added. The absence of the dark blue
colour indicates that the starch has been hydrolyzed and is no longer
available for complexing. Therefore, the lack of blue colour is a positive
result for amylase activity, indicative of the presence of saliva.
3. COLORIMETRIC ASSAY- Phadebas is a synthetic biochemical used for
quantitative assessment of α- amylase. The active component is DSM-P
(Degradable starch microspheres) in which a blue dye chemically bound.
This test is based on the fact that amylase digests starch. The presence
of saliva digests the starch and releases the dye from the complex. The
solution thus becomes blue in color. The degree of coloration is
proportional to the amount of amylase in the sample. This indicates the
presence of saliva. The test can be used as a quantitative test by
measuring the intensity of the developed colour at 620nm wavelength.
4. CONFIRMATORY TESTS
a. Immunochromatographic Assays- Commercially developed
immunochromatographic kits are available for testing saliva samples.
The method is based on antigen-antibody interaction principle. One
such kit is known as RSID saliva kit. The presence of saliva can be
 ascertained by the appearance of pink line in the RSID saliva kit. This
technique can detect saliva as low as 1 µL.
b. RNA based Assays- RNA-based assays have been developed recently
for the identification of saliva. They are based on the expression of
certain genes in certain cell or tissue types. Thus, the techniques
used in the identification of saliva are based on the detection of
specific types of mRNA expressed exclusively in certain cells in the
oral cavity. These assays utilize reverse transcriptase polymerase
chain reaction (RT-PCR) methods to detect gene expression levels of
mRNAs for saliva identification.

Gene symbol Gene product Description

HTN3 Histatin 3 Histidine-rich protein
involved in
nonimmune host
defense in oral cavity
STATH Statherian Inhibitor of
precipitation of
calcium phosphate
salts in oral cavity

Scope :
Saliva is often detected in scenes of crime along with bite marks or lip
prints where the oral cavity may have been involved. Serological and
cellular analysis of thus obtained saliva is of immense use in identification
of the accused. Saliva tests can reveal certain disease markers, viral
infections, and the presence of therapeutic as well as illicit drugs in
the body. Saliva may also contain buccal mucosa cells, it is possible to
identify the DNA profile of the person in question using advanced DNA
profiling techniques.
 Use of saliva in forensics :
SALIVA
(dried saliva stains from bite marks/lip prints)

Comparative identification Reproductive identification

Personal identification SALIVARY SIGNATURE

Human Forensic profile Constituents Advantages

Deoxyribonucleic acid building Geolocation
Salivary •
profiling/fingerprinting • Time of
personal microbiome
collection
characteristics (microbial DNA in
• Personal
saliva) identity
Salivary • Malignancies
biomarkers • Oral & systemic
health status diseases
Salivary • Drug intake
flow/compositio • poisoning
nal changes • alcohol

age Methylation

Hormones level
Gender
like testosterone
Menstrual blood :
➢ Menstrual blood is regular discharge of blood and mucosal tissue (known as
menses) from the inner lining of the uterus through the vagina. This process
begins at what is known as menarche (first cycle) which happens between
12 to 15 years in girls depending on a lot of factors. The process is known as
menstruation and it usually stops (menopause) at the age of 45- 55 years.
➢ Menstrual blood contains sodium, calcium, phosphate, iron, and chloride, the
extent of which depends on the woman. As well as blood, the fluid consists
of cervical mucus, vaginal secretions, and endometrial tissue.
Difference Between Menstrual Blood And Normal Blood :
➢ Menstrual blood is not highly oxygenated; that’s why it is darker than
normal blood. pH is acidic because of the vaginal secretions. The
concentration of Iron, Haemoglobin and protein is found to be less in
menstrual blood than in normal blood in various studies.
➢ Menstrual blood contains vaginal, cervical and decidual cells. It doesnot
coagulate as it is already coagulated. Matrix mettaloproteinases are
present in menstrual blood but not in blood from other sources.
➢ Menstrual blood contains all the five isoenzymes of Lactate
dehydrogenase LDH-1, LDH-2, LDH-3, LDH-4 and LDH-5. Venous blood
contains LDH- 1, 2 and 3 whereas menstrual blood contains all 5.
Examination of menstrual blood :
1. Physical appearance- The discharge is in the form of a trickling descent, it
does not splash. Therefore, the nature and distribution of the suspect stain
must be taken in account when making the examination. Colour
may vary from bright red to dark brown/ black depending on how old it is.
2. Microscopic Examination- Vaginal epithelium and residual cells will usually
be present in menstrual bloodstains, along with endometrial cells.
3. Identification of fibrin degradation product- Menstrual blood is
discharged from the uterus in a fluid state; it contains no fibrinogen and
 cannot be made to clot. This situation is a result of a fibrinolysis process
during which the blood clots subsequently reliquifies prior to menstrual
period. The reliquifaction process is known as fibrinolysis and defined as
enzymatic digestion of fibrin clot to soluble Fibrin Degradation Products
(FDP). Fibrinogen Degradation Products may be assayed by a
haemagglutination inhibition method. In this technique they are quantitated
by their ability to inhibit the agglutination of sheep red blood cells coated
with human fibrinogen by anti- fibrinogen serum.
4. Glycogenated nucleated squamous epithelial cells (cell type found in
vaginal secretion) is detected by Lugol’s iodine assay or periodic acid Schiff
reagent.
5. Detection of Lactate dehydrogenase isoenzyme found in menstrual blood.
6. MMP7 (matrix metalloproteinase); can be found in menstrual blood but not
in circulatory blood; expressed in endometrial tissue, involved in tissue
remodelling during menstruation. It is detected using RT-PCR assay.
Urine :
➢ Urine is the by product of the cellular metabolism in humans. Urine is
yellow-straw coloured fluid produced by the process of urination. It is
excreted out from the body by an opening called urethra. It follows a
pathway from kidney to urinary bladder through ureters.
➢ On average urine production is around 1.4 Litres per day by per person. It
depends from person to person on the various factors like weight, health,
state of hydration and environmental factors.
➢ The normal range for urine pH is 4.5 to 7.8.
Collection & preservation of urine Evidence
➢ In living individual, collect about 100 ml of urine in a sterile container.
➢ Urine in dead maybe collected by direct puncture with needle and
syringe into the bladder after the abdomen has been opened during
 post-mortem examination. It is preserved by adding equal quantity of
saturated common salt solution or rectified spirit or some grains of
thymol.
Forensic importance of urine as evidence :
It can be found in the cases of Strangulation, violent cases, poisoning
cases, sexual assault etc. It can be used to analyse the presence of drugs,
proteins, alcohol and poisons.
Forensic Analysis of urine Evidence :
➢ Visual or physical examination- Locate the stains using alternate light
sources. Observe the colour, clarity and detect any unusual odours. A
suspected urine stain may fluoresce pale yellow or pale blue when
viewed under long and short wave UV light. Safety eyeglasses, which
absorb ultraviolet radiation, must be worn when viewing material for
fluorescence.
➢ Microscopic Analysis- Normal urine contain white blood cells in very
less quantity. Bacteria, yeast cells, parasites are almost negligible.
Abnormal Urine may contain red blood cells in the urine indicative of
kidney or bladder injury, kidney stones or Urinary Tract Infection (UTI).
➢ Chemical Test- Detection of urine stain is based on detection its
constituents like urea, amines, phosphates, sulphates, etc.
1. DETECTION OF UREA (constituent of urine)-
a. Urea in a suspected urine stain can be detected with urease-
bromothymol paper. If present, urea would be enzymatically
decomposed to ammonia and carbon dioxide by urease. This
liberated ammonia will shift the ph. It will change the colour from
yellow to blue.
b. Take the suspected sample on a microscopic slide and add 2-3
drops of concentrated nitric acid. Appearance of rhombohedral
 stacked crystals of urea nitrate which are colourless or
transparent indicates the presence of urine in the sample.
c. Use of urease to catalyse breakdown of urea and release
ammonia and carbon dioxide; ammonia detected using DMAC
(dimethylamino cinnamaldehyde), pink or red colour indicates
the presence of urea in urine.
2. DETECTION OF CREATININE (constituent of urine)-
a. Jaffe colour test- In this test, picric acid is used to convert
creatinine under alkaline condition to form reddish-orange
coloured crystals of creatinine picrate.
b. Colour test with sodium nitroprusside gives Prussian blue colour.
3. DETECTION OF INDICAN- Resorcinol reagent is added to the small
quantity of the extracted stain then cupric bromide solution is added
& mixed and whole mixture is extracted with amyl acetate. The red
colour of the crystal indicates the presence of Indicant.
➢ Confirmatory assays :
1. Urine contains a glycoprotein called Tamm Horsfall protein, produced
exclusively by renal tubular epithelial cells within the distal loop of
Henle. This can be detected by Radioimmunoassay.
2. Thin-layer chromatography is also used to detect both urea and
creatinine in an effort to make the test more specific for urine, and
also additional components of urine on TLC plates can be used for the
same purpose.
❖ Abnormal Constituents of Urine Many a times tests are used in analysing
the urine in order to determine, whether it contains abnormal substances
indicative of diseases or illegal use of some drugs . Usually abnormal
substances that are present in the urine are:
➢ Proteinuria- Protein content in urine, often due to leaky or damaged
glomeruli.
➢ Oliguria- An abnormally small amount of urine, often due to shock or kidney
damage.
➢ Polyuria- An abnormally large amount of urine, often caused by diabetes.
➢ Dysuria- Painful or uncomfortable urination, often from urinary tract
infections.
➢ Haematuria- Red blood cells in urine, from infection or injury.
➢ Glycosuria- Glucose in urine, due to excess plasma glucose in diabetes

SWEAT
➢ Sweat is a fluid secreted by the sweat glands also called sudoriferous
glands. The process of production of sweat is called sweating or
perspiration.
➢ Sweat glands are the exocrine glands which secretes their production
onto epithelial cells by the specific ducts. There are two types of sweat
glands :
1. Apocrine glands- they are mostly present at the armpit area, and
genital areas.
2. Eccrine glands- these glands cover the major surface area of the
body. They are present most in the soles and palms then, the head and
least in the trunk region. These two types of gland differ in a number of
ways, from distribution and structure, to their excretory mechanism and
secretory product.
Structure of a Sweat Gland :
➢ Sweat glands comprise a secretory unit and a duct through which sweat or
secretory product is passed. Sweat glands are situated in the dermis and
are surrounded by adipose tissue. At the base of each sweat gland there is
a structure known as the secretory coil. This is surrounded by contractile
myoepithelial cells which act to help secrete the gland’s product. The
 contractions of these cells are either controlled by hormones or nerve
action.
Examination of Sweat :
➢ Sweat is one of the biological fluids which exhibit the blood group
substances if the person is a secretor, therefore, the analysis of blood
group can be done by examination of sweat. Sweat from secretor individual
will contain the Antigens A, B, and H.
➢ Odour – A small bit from the suspected sample is taken and heated. The
specific odour that comes out is taken note of.
➢ Gee’s urea nitrate test – The sweat bearing areas are first examined for the
presence of urea by the Gee’s urea nitrate test. If the positive result is
found then it is subjected to absorption inhibition, absorption elution or
mixed agglutination test for the identification of blood grouping antigens in
them. Stained material is extracted with acetone. Now, the acetone extract
is set to evaporate to make it more concentrated. The residue is mixed with
acetone solution in a test tube and mixed with glass rod. A slide is prepared
with the drop of solution. Cover it with the cover slips and prevent
overflowing. Leave it for few seconds. Observe the slide under the
microscope. The crystals appear will have following characteristics: long,
colourless, rhombic shaped.
➢ Dermcidin Detection- A monoclonal antibody G 81 has been developed which
react with dermcidin. Using a Western blot, the G-81 antibody only reacted
to the specific peptides in sweat. An antibiotic peptide known as dermcidin
is found to have an identical 18-amino acid segment as the N-terminus of a
peptide that reacted with G-81. ELISA analysis is able to detect the G-81
reactive peptide in sweat samples.
➢ SEM coupled with EDX – It can identify the relative concentrations of
sodium, phosphorus, sulphur, chlorine, potassium, calcium, and other metal
traces. The sweat analysis showed that chlorine and sodium were the only
 consistently clear peaks among the different samples, and potassium was
sometimes visible. The large chlorine peak is used as the basis of
comparison and identification.
Forensic Significance of sweat :
➢ An average square inch of skin contains 650 sweat glands. That means our
bodies leave small amounts of sweat on everything we touch - whether
we’re making a phone call, eating supper or committing a crime.
➢ Sweat stains become important biological evidence in cases like kidnapping,
sexual assault etc. It can also be present on any kind clothing or on victim’s
clothing which can relate us to suspect. It can also be present on surfaces
/ objects touched by victim or suspect.
➢ Sweat can be analysed for determining ethanol, drugs, ions and metals.

Skin :
The skin is the largest organ of the body. The skin and its derivatives (hair,
nails, sweat and oil glands) make up the integumentary system. One of the main
functions of the skin is protection. It protects the body from external factors
such as bacteria, chemicals, and temperature. The skin contains secretions
that can kill bacteria and the pigment melanin provides a chemical pigment
defense against ultraviolet light that can damage skin cells.

Another important function of the skin is body temperature regulation. When

the skin is exposed to a cold temperature, the blood vessels in the dermis
constrict. This allows the blood which is warm, to bypass the skin. The skin then
becomes the temperature of the cold it is exposed to. Body heat is conserved
since the blood vessels are not diverting heat to the skin anymore. Among its
many functions the skin is an incredible organ always protecting the body from
external agents.

Importance :
When a crime scene is searched for evidence, new research indicates that by
swabbing surfaces that were likely touched by the person who committed the
crime, investigators can find an identifiable skin microbial signature the person
left behind. Although the work is preliminary and considered a "first step," one
of the researchers noted that the work has demonstrated that "skin microbes
can ... serve as individual evidence in a variety of scenarios."
Therefore, using skin microbes to identify individuals significantly expands the
scope and power of trace evidence analysis. Our results also reveal two
candidate surface types, plastic and ceramic, to focus technology development
for recovering personalized skin microbial signatures as trace evidence at
crime scenes.
Composition :

Three layers of tissue make up the skin:

1. Epidermis, the top layer of the skin that you can see and touch. Keratin, a
protein inside skin cells, makes up the skin cells and, along with other
proteins, sticks together to form this layer.The epidermis:
• Acts as a protective barrier: The epidermis keeps bacteria and germs
from entering your body and bloodstream and causing infections. It also
protects against rain, sun and other elements.
• Makes new skin: The epidermis continually makes new skin cells. These
new cells replace the approximately 40,000 old skin cells that your body
sheds every day. You have new skin every 30 days.
• Protects your body: Langerhans cells in the epidermis are part of the
body’s immune system. They help fight off germs and infections.
• Provides skin color: The epidermis contains melanin, the pigment that
gives skin its color. The amount of melanin you have determines the
color of your skin, hair and eyes. People who make more melanin have
darker skin and may tan more quickly.
2. Dermis, the middle layer The dermis makes up 90% of skin’s thickness. This
middle layer of skin:
• Has collagen and elastin: Collagen is a protein that makes skin cells
strong and resilient. Another protein found in the dermis, elastin, keeps
skin flexible. It also helps stretched skin regain its shape.
• Grows hair: The roots of hair follicles attach to the dermis.
• Keeps you in touch: Nerves in the dermis tell you when something is too
hot to touch, itchy or super soft. These nerve receptors also help you
feel pain.
• Makes oil: Oil glands in the dermis help keep the skin soft and smooth.
Oil also prevents your skin from absorbing too much water when you
swim or get caught in a rainstorm.
• Produces sweat: Sweat glands in the dermis release sweat through skin
pores. Sweat helps regulate your body temperature.
• Supplies blood: Blood vessels in the dermis provide nutrients to the
epidermis, keeping the skin layers healthy.
3. Hypodermis, the bottom or fatty layer.The bottom layer of skin, or
hypodermis, is the fatty layer. The hypodermis:
• Cushions muscles and bones: Fat in the hypodermis protects muscles
and bones from injuries when you fall or are in an accident.
• Has connective tissue: This tissue connects layers of skin to muscles
and bones.
• Helps the nerves and blood vessels: Nerves and blood vessels in the
dermis (middle layer) get larger in the hypodermis. These nerves and
blood vessels branch out to connect the hypodermis to the rest of the
body.
• Regulates body temperature: Fat in the hypodermis keeps you from
getting too cold or hot.
One inch of your skin has approximately 19 million skin cells and 60,000
melanocytes (cells that make melanin or skin pigment). It also contains 1,000
nerve endings and 20 blood vessels.
Scope :
Skin detection is process of extracting potential skin color regions from an
image or video. Skin color detection restricts the work-space as it showcases
human face, limbs and other exposed body features out of the whole image.
Thus it can be used as a processing step in variety of applications like face
detection and face recognition. Extraction of skin color pixels also help to
identify pornographic image from the database of images and efficiently used
for forensic purpose. It is been observed that there is positive correlation
between images having large skin regions and the images which are
pornographic in nature. We have implemented this approach using filtering of
images in RGB and YCbCr color spaces using specific threshold values.

Nails :
Nail is one of the physical evidences that can be encountered in criminal cases
related to homicide and sexual offences etc. Though it is not widely found on
the crime scene but if at all encounters it should not be avoided or destroyed
as it can help us to narrow down our area of search.
Unlike other type of biological evidence, nail is remarkably stable evidence to
most of the environmental conditions and does not break down easily. In
addition it is relatively unnoticeable to the untrained eye; therefore a criminal
is not likely to make special efforts to destroy the nail evidence, as it is not
known to many people that this small nail clipping can also play important role
in linking the criminal to either to crime scene or a victim.
Evaluation of human nail clippings provides useful information which helps us in
achieving the individuality of an individual. In the present study attempt has
been made to study the microscopic features of nail striations present on the
nails with the help of comparison microscope. In this study the parameters
that were included were, matching of inter finger and toe nail striations and
intra finger and toe nail striation also the nail striations of different individuals
were matched.
nail as evidence may be used to:
1. To study the Uniqueness in nail striations
2. To study Permanency and persistence in nail striations

Tissue :
A tissue is a series of cells that accomplish a shared function. Tissues, in
turn, form organs, such as the stomach and kidney.
Body tissue identification are important in forensic science as they can provide
key evidence in a criminal investigation and may assist the court in reaching
conclusions. Establishing a link between identifying the fluid or tissue and the
DNA profile adds further weight to this evidence.

Tooth :
➢ Just like your fingerprints or snowflakes, there are no two teeth that are
alike. Your teeth are unique and of one kind. Not even identical twins have
the same teeth. Since teeth are specific to every individual they are often
used as an identifier.
Significance of Teeth In Forensic
➢ The external layer of teeth is composed of enamel which is the hardest
tissue of the body so it can hold out against trauma, decomposition, heat,
degradation, water immersion, desiccation better than other body tissues.
➢ Teeth are the most durable organ in the body and can be heated to a
temperature of 1600 degrees Celsius without appreciable loss of
 microstructure and are usually the only remains after an extended period
of burial. Teeth are an outstanding source of DNA since they can stand up
against conditions like humidity, temperature, and microbial action.
Purpose of Forensic Odontology
• Forensic odontology plays a major role in both death investigations as well
as in the evaluation of victims of sexual assault and child abuse. The dental
practitioner helps by keeping accurate dental records and providing all
necessary information and identify individuals
• Human dentition consists of incisor, canine, premolar, and molar that has a
various dental feature like morphology, variations in shape and size,
restoration, pathologies, missing tooth, wear pattern, crowding, color,
position, rotation, dental anomalies which gives every individual unique
identity.
• Species Identification- Based on the morphology and anatomy of the tooth it
can be identified whether it is human or not. It is possible to determine even
by dental tissue because dentinal fluid contains special information that can
be compared and analyzed using counter current electrophoresis with
artificial antisera.
• Race Determination- Human species have been categorized into three races
Caucasoid , mongoloid , negroid. Dental characteristics such as shoveling,
taurodontism, carabelli's cusp hypocone, protostylid, peg shaped incisor,
etc are used to determine ancestry. Dental restoration also helps in finding
the ethnicity of an individual. The type of dental treatment recommends the
financial status of a person. Teeth provide important evidence regarding the
habits and occupation of the individual.
• Gender Determination- Sex determination plays a major part in the
recognizable proof of unknown persons in a natural calamity, chemical, or
nuclear bomb explosion. It can be done using odontometric techniques
which are measurements of the tooth. Dental indices are used to show
sexual dimorphism. Gender can also be determined by the presence of sex
chromatin or Barr bodies in the pulp.
• Age Estimation- The requirement for age estimation has increased because
of growth in the number of unidentified cadavers and human remains and
cases for living people with no substantial confirmation of date of birth. The
number and sequence of teeth erupted in the jaw help in the determination
of age. The radiographic method elaborates on various stages of
mineralization and helps in giving an accurate estimation of age.
• Bitemark Analysis- ABFO defines bitemarks as patterns left in an object or
tissue by the dental structure of an animal or human. Bitemark inflicted by
teeth are considered to be highly individualistic to a person and has been
used since Roman times. Teeth are used as a weapon by an aggressor and
as self-defense by the victim. Peculiar characteristics like missing teeth,
malformed teeth, fractured teeth, crowding, diastema are helpful in the
comparison process.
• Comparative Dental Identification- Teeth play a critical role when the
recognizable proof of remains of a perished individual is skeletonized,
deteriorated, burned, or dismembered and cannot be distinguished by visual
or unique strategies. The core of the identification procedure is comparing
the post mortem remains with antemortem records which include notes,
radiographs, study cast, etc. In Disaster Victim Identification the dental
evidence collected is compared to the antemortem records available to the
dentist for identification of a deceased person.
• DNA Analysis- Teeth are fabulous source of genomic DNA due to its
resistant nature. In teeth, DNA is found in vascular pulp, odontoblastic
process, accessory canal, and cellular cementum. Even the root-filled teeth
provide sufficient material for PCR analysis which produces a DNA profile
which is further compared with antemortem samples or paternal DNA
Bones :
Bone is a mineralized connective tissue from which bones, the main component
of the vertebrate skeleton, are formed. Bone tissue is composed of cells,
namely osteoblasts, osteoclasts and osteocytes, and an extracellular matrix
comprising inorganic hydroxyapatite crystals and organic collagen.
Importance :
bones are visually, stereoscopically, and radiographically examined so that we
can determine the age, sex, stature, and ancestry of the victim.
Identification can be done by studying the bones of the human corpse during
autopsy examination and if unknown skeletal remains are all that is available,
examination of each bone is required.
Bones marked by perimortem injuries, such as unhealed fractures, bullet holes,
or cuts, can reveal cause of death
Bones contain information about people's lives such as where they came from,
their age at death and which diseases they suffered from.
Bone classification :
Bone
Features Function(s) Examples
classification

Femur, tibia, fibula,

metatarsals,
Cylinder-like
Movement, humerus, ulna,
Long shape, longer than
support radius,
it is wide
metacarpals,
phalanges

Cube-like shape,
Provide stability,
approximately
support, while
Short equal in length, Carpals, tarsals
allowing for some
width, and
motion
thickness

Points of
attachment for Sternum, ribs,
Flat Thin and curved muscles; scapulae, cranial
protectors of bones
internal organs

Vertebrae, facial
Irregular Complex shape Protect internal
organs, bones
 Bone
Features Function(s) Examples
classification

movement,
support

Protect tendons
Small and round; from excessive
Sesamoid embedded in forces, allow Patellae
tendons effective
muscle action

Scope :
scientists be able to determine from a whole bone or part of a bone are
1. an age range
2. sex
3. race
4. approximate height
5. cause of death, disease, or anomaly

Uterine fluid :
Before and during embryo implantation, uterine fluid is the liquid medium that
connects the floating embryo(s) and the uterus, and has the potential to
transfer vital information between the embryo and the uterus.
Determination of the type and origin of the body fluids found at a crime scene
can give important insights into crime scene reconstruction by supporting a
link between sample donors and actual criminal acts. For more than a century,
numerous types of body fluid identification methods have been developed, such
as chemical tests, immunological tests, protein catalytic activity tests,
spectroscopic methods and microscopy. However, these conventional body
fluid identification methods are mostly presumptive, and are carried out for
only one body fluid at a time. Therefore, the use of a molecular genetics-based
approach using RNA profiling or DNA methylation detection has been recently
proposed to supplant conventional body fluid identification methods.
Several RNA markers and tDMRs (tissue-specific differentially methylated
regions) which are specific to forensically relevant body fluids have been
identified, and their specificities and sensitivities have been tested using
various samples. In this review, we provide an overview of the present
knowledge and the most recent developments in forensic body fluid
identification and discuss its possible practical application to forensic
casework.

Vomit :
➢ Vomiting is the involuntary, forceful expulsion of the contents of one’s
stomach through the mouth and sometimes the nose. Emesis is the medical
term for vomiting.
➢ The pH of the vomitus is almost always highly acidic and is associated with a
foul smell.
➢ Sometimes the vomitus may be streaked in blood and be a red and brown
colour. This is called blood vomiting or hematemesis. Fresh blood in the
vomit appears red and usually comes from the upper gastrointestinal tract.
Blood that comes from the lower gastrointestinal tract usually undergoes
oxidation and may appear brown in color. Clotted blood appears dark red in
color and is usually seen when there is perforation of a peptic ulcer.
Contractions of the duodenum lead to the secretion of bile in the vomitus.
This gives a greenish tinge to the vomit. Severe vomiting may often yield
 green coloured vomit, although a bile-containing vomitus may also be yellow
in colour.
Examination of vomit :
1. Test for Mucus- To the extract add 33% acetic acid drop by drop.
Opalescence appears which may be due to mucus or lipoid substance or
both. If on addition of more acetic acid opalescent does not disappear,
presence of mucus is confirmed, because with excess of acetic acid lipoid
globulins dissolve but not the mucus.
2. Test for Free Hcl (Gunzberg’s Test)- The reagent is prepared by 6 drops of
10% phloroglucinol in alcohol with 3 drops 10% vanillin in alcohol.In a
porcelain evaporating dish one drop of suspected extract is placed and 1-2
drops of Gunzberg’s reagent is mixed at once. The contents are allowed to
dry completely. A brilliant red colour indicates free HCl.
3. Endothelial cells- After centrifuging the extract for 10 minutes a thin film is
made on a slide. The Endothelial Cells are observed under microscope. 4.
ELISA Detection of Gastric Mucosa-Expressing Proteins- Four gastric
mucosa-expressing proteins, pepsinogen I (PGA), pepsinogen II (PGC),
gastrin (GAST), and mucin 5AC (MUC5AC) are used to detect the presence of
vomit. Enzyme-linked immunosorbent assay (ELISA) procedure is used for
the detection of these four proteins.
Forensic Significance of Vomit :
The identification of vomit stains may be helpful for crime scene
reconstruction. Vomitus can be found in the cases of poisoning or drug abuse,
hanging or sexual abuse. The questioned sample could be tested for the
presence or absence of any particular drug or poison if administered. This
evidence is very rarely found at the crime scene and is not much evidentiary

Vitreous humour :
➢ Vitreous humor is the fluid-like gel, composed of approximately 98–99%
water with trace amounts of hyaluronic acid, glucose, anions, cations, ions,
and collagen, located in the posterior chambers of the eyes (about 3 ml per
eye).
➢ Vitreous humor is useful for the postmortem measurement of alcohol
because it remains sterile for days after death and is therefore not subject
to artifactual alcohol formation (unlike postmortem blood). Due to the
higher water content of vitreous compared to whole blood, at equilibrium,
vitreous alcohol concentrations are typically 10–15% higher.
➢ Vitreous fluid has also been used for the measurement of drugs. However,
its use is limited by two factors. First, the volume available is usually limited
to about 6 ml, even if both eyes are used. The second factor is that vitreous
drug concentrations can be very different from the corresponding whole
blood or plasma concentrations. Concentrations of highly lipid soluble or
highly protein-bound drugs tend to be much lower in vitreous fluid, due to
the relative absence of lipid material and much lower protein content than
whole blood.
➢ As the vitreous humor is isolated by membranes from the remainder of the
decomposing body, attempts have been made to use the chemistry of the
fluid to estimate the postmortem interval. The concentrations of both
potassium ions and hypoxanthine have been used. Vitreous potassium
increases in concentration after death. Hypoxanthine is a degradation
product of adenosine and increases due to hypoxia before death.
➢ Vitreous humor is preferred to blood for most postmortem biochemistry
since it is thought far less susceptible to autolytic change, is less likely to
be subject to postmortem contamination by diffusion of drugs or other
poisons that may be present at high concentration in the thorax or abdomen
at death, and lies within the relatively protected environment of the eye
socket.

CSF :
cerebrospinal fluid The fluid that flows in and around the hollow spaces of the
brain and spinal cord, and between two of the meninges (the thin layers of
tissue that cover and protect the brain and spinal cord). Cerebrospinal fluid is
made by tissue called the choroid plexus in the ventricles (hollow spaces) in
the brain. Also called CSF.
CSF acts as a cushion, protecting the brain and spine from injury. The fluid is
normally clear. It has the same consistency as water. The test is also used to
measure pressure in the spinal fluid.
The purpose of a CSF analysis is to diagnose medical disorders that affect the
central nervous system. Some of these conditions include:

• viral and bacterial infections, such as meningitis, West Nile virus, herpes vi
rus, and encephalitis
• tumours or cancers of the nervous system
• syphilis, a sexually transmitted disease
• bleeding (haemorrhaging) around the brain and spinal cord
• multiple sclerosis, a disease that affects the myelin coating of the nerve fib
ers of the brain and spinal cord
• Guillain-Barr syndrome, an inflammation of the nerves.
• Early onset Alzheimer’s disease the levels of two substances known as
amyloid beta (1-42) and phosphorylated tau in CSF appear to be useful
diagnostic markers for earl y-onset Alzheimer’s.
• CSF analysis is also used in forensic investigations to identify the presence
of illicit drugs (e. g., heroin) or poisons in the bodies of murder, accidental o
verdose, or suicide victims.

Collection and preservation :

CSF protects the brain and spinal cord from injury by acting like a liquid
cushion. CSF is usually obtained through a lumbar puncture (spinal tap). During
the procedure, a needle is inserted usually between the 3rd and 4th lumbar
vertebrae and the CSF fluid is collected for testing
Use a sterile syringe and needle to inoculate 0.5-1.0 ml of CSF into the T-I
medium. The remaining CSF should be kept in the collection tube. It should not
be refrigerated, but should be maintained at room temperature (20-25°C)
before Gram staining and other tests.

Colostrum :
Colostrum is the first form of breastmilk that is released by the mammary
glands after giving birth. It's nutrient-dense and high in antibodies and
antioxidants to build a new born baby's immune system. It changes to breast
milk within two to four days after your baby is born.

Botanical materials :
Forensic botany is the scientific use of plant materials to help solve crimes. It
is study of plant life in order to gain information regarding possible crimes.
Forensic botany is primarily engaged in making connections between evidence
and a crime. Various plants or plant materials such as pollen at the scene of
crime or a rare plant type present near a murder victim can be helpful in
connecting a suspect to a victim or scene.
Forensic botany is similar in its role to that DNA FINGERPRINTING. just as
fingerprint are unique to individuals, plant material is often unique to certain
ecological areas or plant species
Plants allow forensic botanist to identify things such as what season the crime
took place, or geographical location, whether a body has been moved following
a murder& how long a body has been buried if it was buried. Plant materials
can be found on dead bodies, clothes, floor etc.
Homicide or Suicide: These cases were reported in Taiwan which shows the
simple use of plants. The first case, a body of a young woman was found in the
gutter. Since no bone fractures were seen (before the autopsy) it was taken as
a hit and run case. On viewing the surveillance tapes she was seen but as soon
as a truck passed her she was not seen so it was concluded that she has been
hit by the truck and to hide the accident the driver has hid her body in the
gutter. While during the autopsy some plant material, a tiny berry and stem,
were found in her hairs which were uncommon for that area suspected to be
from the family Solanaceae. Later on, while the investigators visited the scene
of crime they found the similar plants on the railing of the building present
there whose height was such that no person walking on street can come into
its contact. Thus they concluded that the woman has felled from the top of
building and while falling down her body made the contact with the plants and
so the plant materials were transferred to her hairs and felled into the gutter.
The autopsy report mentioned the cause of death as the impact injuries and the
interviews held with her relatives confirmed her suicidal tendency due to her
suffering from depression.
Forensic botany or the uses of plants in criminal investigation their primary
role in criminal investigation is in making connections between evidence &
crime Fibers, hair, soil, wood, gunshot residue and pollen are only a few
examples of trace evidence that may be transferred between people,objects or
the environment during a crime. Investigators can potentially link a suspect
and a victim to a mutual location called as trace evidences ,The materials
which can be seen from naked eye without the use of magnifying glass are
called micro botanicals
determine the circumstances &cause of death
 Estimate time frame in relation to the death
 Establish where the death could have taken place
 Determine if there were multiple crime scene
 Prove or disapprove an alibi
 Place the suspect at the scene of crime
One of the particular application of counting annual ring in roots help in
determining When a body was buried for example the ground around where
a body was found has been shown to e disrupted it can be assumed that any
new growth began approximately by counting the number of rings in a new
growth a date within a year can be acquired If a plant that is in the place
whre a body is buried is damage it is infact possible to count the number of
 annual rings that have een formed since the injury by studing the length &
size of roots found in burial state an investigators can rely on independent
data that approxiamete average root growth for the same plant found at the
grave. In the case of stem if two pieces of stem found from a tree to know
about whether they belong to the same tree they can be identified on the
basis of external contour of the stem
plant essential features have become increasingly more common in
forensic application A record what the victim ate just before the death can
be obtained by identifying plant cells peresent in stomact content. In some
exact restronaunt that served the last meal can be estimated plant cells
and their associated hair or trichomes were able to be identified because of
their relative resistance of cellulose wall against gastric juice  Flower
morphology gives an opportunity to identify floral species using a
dichotomous key. Correct identification of the floral species will help to
determine the location of a recent murder, giving police valuable
information as to the identity of the suspect.

Diatoms :
Diatoms are photosynthesising algae, they have a siliceous skeleton
(frustule) and are found in almost every aquatic environment including fresh
and marine waters, soils, in fact almost anywhere moist.
Diatoms have been used in forensic science in a variety of ways, the most
frequent being the diagnosis of death by drowning. When a person drowns,
water will enter the lungs and then enter the bloodstream through ruptures in
the peripheral alveoli before being carried to the other organs such as the
liver and heart.
Diatoms are unicellular eukaryotic microalgae that play important ecological
roles on a global scale. Diatoms are responsible for 20% of global carbon
fixation and 40% of marine primary productivity. Thus they are major
contributors to climate change processes, and form a substantial basis of the
marine food web.

Wild life samples :

wildlife forensics is a field of criminal investigation wherein science is used to
identify and examine evidence from crime scenes where animals have been
killed, particularly those that are protected by law. Wildlife forensics plays a
crucial role in curtailing the wildlife trade and human-wildlife conflict.
Wildlife helps in maintaining the eco-logical balance of nature. Killing of
carnivores leads to an increase in the number of herbivores which in turn
affect the forest vegetation, thus due to lack of food in the forest they come
out from the forest to agriculture land and destroy our crops.
Wildlife forensics is concerned with providing scientific evidence to inform
investigations into crimes against wildlife, focusing on determining the identity
of poached or illegally traded wildlife products, and addressing questions
relating to the species, geographic origin, relatedness, individual identity and
age of samples.

HAIR : HAIR TRICHOLOGY

➢ Hair is a protein filament that grows from follicles found in the dermis.
➢ Hair is one of the defining characteristics of mammals.
➢ The hair is made up of 95% keratin, a fibrous, helicoidal protein (shaped like
a helix) that forms part of the skin and all its appendages (body hair, nails,
etc.). Keratin is synthesized by keratinocytes and is insoluble in water, thus
ensuring impermeability and protection for the hair.
➢ Some 18 amino acids can be found in the hair, such as proline, threonine,
leucine and arginine. Keratin is particularly rich in cysteine (a type of
 sulfurated amino acid), which forms disulfide bonds between molecules,
adding rigidity and resistance to the entire structure.
➢ The study of hair is known as trichology.
Structure of Hair :
The word "hair" usually refers to two distinct structures:
➢ The part beneath the skin, called the hair follicle, or, when pulled from the
skin, the bulb or root. This organ is located in the dermis and maintains
stem cells, which not only re-grow the hair after it falls out, but also are
recruited to regrow skin after a wound.
➢ The hair shaft, which is the hard filamentous part that extends above the
skin surface.
The hair shaft is made up of three layers
1. Cuticle- It is the outermost layer. It comprises large number of mostly
transparent and overlapping scales. The distal part of each scale usually
covers the proximal part of the next scale. Animal hairs have scale patterns
that vary by species, and these patterns are a useful diagnostic tool for
identifying animal hairs. Humans have a scale pattern called imbricate, but
it is fairly common among animals and, despite attempts to use scales as an
individualizing tool for human hairs, is not generally useful in forensic
examinations.
2. Cortex- It is the middle layer. It comprises dead cornified cells that are
packed into rigid and homogenous mass. Cellular details cannot be identified
in the cortex due to high cornification and close packing and as such, cortex
do not have much value in forensic. In human hair, cortex is the largest part
and contains The granules vary in size, shape, aggregation, and distribution
excellent characteristics for forensic comparisons. Small bubbles, called
cortical fusi, may appear in the cortex; when they do appear, the may be
sparse, aggregated, or evenly distributed throughout the cortex. Cortical
fusi also vary in size and shape.
3. Medulla- It is the innermost layer. It can be hollow or may be comprised of
shrunken dead cells with spaces filled with air. Medulla is very valuable in
species identification. 5 different groups depending on appearance of
medulla-continuous, fragmented, interrupted, solid and none.

Biology of Hair :
Hairs grow from the skin, or, more precisely, the epidermis, of the body. The
follicle is the structure within which hairs grow; it is a roughly cylindrical tube
with a larger pit at the bottom. Hairs grow from the base of the follicle
upwards. In the base of the follicle, the hair is still very soft; as the hair
proceeds up the follicle, it slowly begins to harden and dry out. Hair is made of
keratin, a tough, protein-based material that makes hair, nails, and horns in
animals. The hardening process of hair growth is of the most durable materials
produced by nature and hairs from mummies, both natural and cultural in
origin, have been found thousands of years after the person’s death.
Keratinisation also explains why it does not hurt when a person’s hair is cut:
Hair is “dead” from the moment it peeks above the skin. The only place hair is
“alive” is in the base of the follicle, which is why it hurt when a hair gets pulled
out. The follicle contains other structures, such as blood vessels, nerves,
sebaceous gland the last producing oils that coat our hairs, helping to keep
them soft and pliable. Hairs even have muscles, called pili arrector muscles
(pilus is the Latin word for “hair”), which raise hairs when a person gets chilled
(so-called goose bumps). Specialized cells in the follicle produce small colored
granules, called melanin or pigment, that give hairs their particular color;
these cells are called melanocytes. Only two types of melanin are found in
hairs: a dark brown pigment called eumelanin and a lighter pigment called
phaeomelanin. The combination, density, and distribution of these granules
produce the range of hair colors seen in humans and animals.

Examination of Hair :
1. Determination of AGE - Age can be determined sometimes from hair, but
within wide limits, as between an infant and an adult. Roots of hair from
children will dissolve rapidly in a solution of caustic potash, but in older
 people roots will resists this treatment. Age can also be determined by the
type of hair- lanugo, vellus and terminal hair. Lanugo hairs are formed in
the embryo as the first product of follicular development. These hairs are
fine and soft, usually not medullated nor pigmented. They are replaced by
vellus hairs which appear just before birth. Terminal hairs represent the
final state of differentiation in humans. They are long, pigmented, coarse
and sometimes with medulla. Terminal hair is of two types- primary
terminal hairs which appear at scalp, eyebrows, eyelashes etc. another one
is secondary terminal hair which develop at puberty such as axilla and pubic
hair.
2. Determination of SEX - studying the sex chromatin (X & Y bodies) from hair
root cells of the scalp. In addition, beard and moustache hairs of the males
are the only hair whose sex can be determined. Male hairs are generally
coarser, thicker and darker as compared to females. In human head hair,
Barr bodies are found in the hair follicles in a proportion of 29±5 percent in
females and 6±2 percent in males.
3. Determination of SOURCE (From Which Part Of The Body)- Unlike other
animals, humans exhibit a wide variety of hairs on their bodies. The
characteristics of these hairs may allow for an estimation of body area
origin. The typical body areas that can be determined are head (or scalp),
pubic, facial, chest, axillary (armpits), eyelash/eyebrow, and limb. Hairs that
do not fit into these categories may be called transitional the chest and the
pubic region.
4. Determination of Ethnic ORIGIN - The morphology and colour of a hair lying
on a surface can give an indication of a person’s ancestry. Humans are
more variable from one to another in their hair morphology than any other
primate. This variation tends to correlate with a person’s not an exact
correlation. groups are used: Europeans, Africans, and Asia hairs provide
the clearest evidence for ancestral estimates. It may be possible with
certain other hairs, especially facial hairs, but body hairs should be viewed
cautiously.
5. DNA Profiling- Forensic hair analysis can be criminal by evaluating hair
structure and DNA from cells attached to the root of the hair.

Fiber : Fiber is a type of carbohydrate that the body can't digest. Though
most carbohydrates are broken down into sugar molecules called glucose,
fiber cannot be broken down into sugar molecules, and instead it passes
through the body undigested.
Types of fiber :
1. Natural fiber
2. Synthetic fiber
‘Natural fibre’ is a term used to refer to the fibres that are obtained from (or
are produced by) animals and plants. These fibres have a wide range of
applications in the manufacture of composite materials. Paper and felt (a type
of textile material) can be prepared by matting different layers of natural
fibres into sheets. Eg : plant, animal and mineral

➢ Plant Fibers

• Seed fibres – the fibres obtained from the seeds of different types of
plants.
• Leaf fibres – the natural fibres that can be collected from the leaves of
certain plants. Examples include pineapple and banana leaf fibres.
• Fruit fibres – the natural fibres that are obtained from the fruit of a
plant (coconut fibre, for example).
• Stalk fibres – the natural fibres that are obtained from the stalks of
certain kinds of plants. Examples include the wheat straws, bamboo
fibres, fibres obtained from the stalk of rice and barley plants, and
straw.
• Bast fibres – the natural fibres that are obtained from the cells
belonging to the outer layer of the stem. Examples of bast fibres include
jute fibres, flax fibres, vine fibres, industrial hemp fibres, kenaf fibres,
rattan fibres, and ramie fibres. It can be noted that these fibres are
widely used in fabric and packaging due to their durable nature.

➢ Animal Fibres
• Animal fibres are natural fibres which usually contain proteins like
fibroin, keratin, and collagen. Common examples on animal fibres are
listed below.
• Silk – animal fibres that are obtained from silkworms (different species
produce different types of silk).
• Sinew – animal fibre that connects the muscles of certain animals to
their bones.
• Wool – animal fibre that is obtained by shearing off the fur of certain
breeds of sheep.
• Mohair – animal fibre crafted from the hair of the Angora goat.
➢ Mineral fibers
• chemical and Physical Data. 'Man-made mineral fibres' is a generic term
that denotes fibrous inorganic substances made primarily from
rock, clay, slag or glass. These fibres can be classified into three
 general groups: glass fibres (comprising glasswool and glass filament),
rockwool and slagwool, and ceramic fibres.
• Examples of mineral fibers are Asbestos, graphite, and glass

Synthetic fibres are man-made fibres produced from chemical substances and
are used for making clothes and other useful things. These are made by the
process of polymerization. Synthetic fibres can either be completely synthetic
or semisynthetic. Fibres that are purely synthetic like nylons, polyesters,
acrylics are made from chemicals whereas semisynthetic fibres such as
rayons are produced with the utilization of natural polymers as raw material.
Synthetic fibres are of four types, namely: Rayon, Nylon, Polyester, Acrylic

➢ Rayon

• Rayon has properties similar to those of silk.

• It is a man-made fibre and cheaper than silk.
• It is obtained from wood pulp.
• It is infused with cotton or wool to prepare bedsheets and carpets
respectively.
• It is also known as artificial silk and can be dyed in a wide variety of
colours.

➢ Nylon

• These are strong elastic and light, lustrous and easy to wash fibres
made from water, coal, and air initially.
• The fibre is completely synthetic and stronger than steel wire.
• It is used to make socks, ropes, toothbrushes, tents, seat belts,
curtains, etc.
• Used to make ropes for rock climbing and parachutes.

➢ Polyester

• Polyester is made up of many units of an ester.

• It is suitable for making dress material because it is easy to wash and
stays crisp and wrinkle-free.
• Terylene is a known polyester.
• PET (Polyethylene terephthalate) is used to make many useful products
like bottles, utensils, films, wires.

➢ Acrylic

• Many sweaters and blankets are not created from natural wool but from
a kind of synthetic fibre known as acrylic.
• The clothes prepared from acrylic are cheaper and more durable.
• Acrylic is more prevalent than natural wool.
 • However, synthetic fibre melt on heating. If they catch fire, it could be
really dangerous. The fabric sticks to the body of the person wearing it.
Therefore, one should avoid acrylic clothes while in the kitchen or a
laboratory.

Fibre analysis :
• Fibres are useful as a trace evidence in the investigation process as the
origin of fibres can be identified.
• Fibres are thread-like elements from fabric or textile materials. Fibre
evidence can be considered as circumstantial evidence and strengthen any
additional circumstantial or solid evidence against a culprit.
Collection techniques for fibre evidence :
• Picking: Trace evidence like fibre can be picked up by using a clean
forceps.
 • Lifting: In this method, an adhesive bearing substrate such as tape is
used for the collection of trace evidence.
• Vacuum Sweeping: In some cases, a vacuum cleaner along with the filter
trap is used to recover trace evidence from an item or area.
• Clipping: Trace evidence present in fingernail can be recovered from
clipping. Clean clippers or scissors are used to clip fingernails.
Forensic analysis of fiber evidence
1. Preliminary Examination- The fibres are examined visually with a hand
magnifier and under stereomicroscope. The study includes: The twist of the
thread, string, rope or cord.
• The number of strands
• The number of threads in the string
• The number of fibres in each thread.
• the defect in the thread or weave pattern.
• If recovered torn piece of cloth originally formed part of the standard
provided, a mechanical fit may indicate the common source.
2. Microscopic Examination- Microscopic examination is useful to find:
• The structure of fibre
• The nature of fibre materials
• The diameter of the fibre
• The presence or absence of contamination
• Color of fibre
• Cross sectional structure
3. Physical Examination- Density, refractive index, melting and softening point
and tensile strength give important information about the fibres.
4. Fiber Colour- The value of fibre identification is influenced by color. Various
dyes are used to give fibre its desired color. Application of color and
absorption along the length of the fibre are important comparison
characteristics. Discoloration and color fading also leads to increased value of
fibre association. The dyes of crime and specimen samples can be compared
using comparison microscope and other instrumental techniques.
5. Solubility Test- Test the solubility and decomposition of a fabric using strong
acids (Hydrochloric acid or sulfuric acid) or strong bases (NaOCl, acetone,
NaOH).This determines the fabrics polymers.
6. Burn test- Look at how a fiber burns, its odor, and appearance of ash.
Leather :
Leather is a strong, flexible and durable material obtained from the tanning, or
chemical treatment, of animal skins and hides to prevent decay. The most
common leathers come from cattle, sheep, goats, equine animals, buffalo, pigs
and hogs, and aquatic animals such as seals and alligators.
Animal leather :
Today, most leather is made of cattle hides, which constitute about 65% of all
leather produced. Other animals that are used include sheep (about 13%),
goats (about 11%), and pigs (about 10%). Obtaining accurate figures from
around the world is difficult, especially for areas where the skin may be
eaten.Other animals mentioned below only constitute a fraction of a percent of
total leather production.
Horse hides are used to make particularly durable leathers. Shell cordovan is a
horse leather made not from the outer skin but from an under layer, found only
in equine species, called the shell. It is prized for its mirror-like finish and anti-
creasing properties.
Lamb and deerskin are used for soft leather in more expensive apparel.
Deerskin is widely used in work gloves and indoor shoes.
Reptilian skins, such as alligator, crocodile, and snake, are noted for their
distinct patterns that reflect the scales of their species. This has led to hunting
and farming of these species in part for their skins. The Argentine black and
white tegu is one of the most exploited reptile species in the world in the
leather trade. However, it is not endangered and while monitored, trade is legal
in most South American countries.
Kangaroo leather is used to make items that must be strong and flexible. It is
the material most commonly used in bullwhips. Some motorcyclists favor
kangaroo leather for motorcycle leathers because of its light weight and
abrasion resistance. Kangaroo leather is also used for falconry jesses, soccer
footwear,(e.g. Adidas Copa Mundial) and boxing speed bags.
Although originally raised for their feathers in the 19th century, ostriches are
now more popular for both meat and leather. Ostrich leather has a
characteristic “goose bump” look because of the large follicles where the
feathers grew. Different processes produce different finishes for many
applications, including upholstery, footwear, automotive products, accessories,
and clothing.
In Thailand, stingray leather is used in wallets and belts. Stingray leather is
tough and durable. The leather is often dyed black and covered with tiny round
bumps in the natural pattern of the back ridge of an animal. These bumps are
then usually dyed white to highlight the decoration. Stingray rawhide is also
used as grips on Chinese swords, Scottish basket hilted swords, and Japanese
katanas. Stingray leather is also used for high abrasion areas in motorcycle
racing leathers (especially in gloves, where its high abrasion resistance helps
prevent wear through in the event of an accident).
Leather from endangered species :
Exotic leathers are either made from relatively rare animal species or from
skin parts of animals that are rarely processed into leather. Some exotic
leathers are protected by the CITES (Convention on International Trade in
Endangered Species of wild fauna and flora). Crocodile or snake skin leathers
are commonly addressed as leather. However, other exotic leathers like fish
leather, leather from chicken legs or from cow belly/stomach also exist.
Depending on the culture, the definition of exotic leather, can differ. Some
exotic leather surfaces may feel softer, hence making them more appealing, or
they might have a very different look. Fur for instance can be particularly soft
or have a unique colour. However, demand for leather is generally satisfied by
the hides from animals that account for 90% of world meat production. Exotic
leathers are unique mainly because of their texture, look and colour which
makes them more valuable.
Artificial leather :
Artificial leather, also called synthetic leather, is a material intended to
substitute for leather in upholstery, clothing, footwear, and other uses where a
leather-like finish is desired but the actual material is cost prohibitive or
unsuitable. Artificial leather is known under many names, including leatherette,
imitation leather, faux leather, vegan leather, PU leather, and pleather
Paper :
Paper is a thin sheet usually manufactured from cellulose pulp derived from
wood and other lignocellulosic materials such as cotton, rice or wheat straw
for writing, printing and packaging purposes.
The first step in a typical paper manufacturing process is to produce pulp from
wood chip. Softwoods, such as spruce and pine with slender, strong and elastic
fibers
Paper, can be manufactured using two methods: chemical pulping or
mechanical pulping.
The first one involves breaking down the chemical structure of lignin into a
liquid using different chemicals, including sodium hydroxide and sodium sulfide.
Cooking liquor is a by-product of the production, which is washed from
cellulose fibers to produce pulp. Chemical pulping is used to produce higher
quality paper with more expensive production cost than that of mechanical
pulping.

Mechanical pulping can further be classified into two subgroups, namely

ground pulping and thermo-mechanical pulp (TMP) that does not remove lignin
from the fibers in contrast to chemical pulping. In both methods chips are
simply fed into a refiner to disintegrate and to convert the material into fiber
bundles. The refiners are consisted of steam-heated rotating steel discs having
different types of profiles. The final product of TMP is unbleached, dark pulp
with short-length fibers. The main advantage of this kind of pulp is it has a
higher yield than that of chemical pulping. The second one is a more commonly
used method to produce paper with low strength properties.
For writing purposes whiteness of paper is important, therefore, pulp is
bleached using mostly oxygen bleaching techniques rather than chlorine
bleaching due to its high environmental pollution problem. Dark color lignin is
removed during the bleaching process. Most of the strength of paper comes
from hydrogen bond between fibers. Beating and refining of the pulp increase
surface area of fibers so that better contact between fibers will result in
higher mechanical properties of the paper.

A conical refiner is a widely used machine to improve pulp quality

Pulp flows on the screen of the Fourdrinier, and water is drained away with the
help of a series of vacuum boxes and other equipment before a thin sheet of
fiber mat is formed. Speed of the sheet in the machine ranges from 1,200 fpm
(13.6 mph) to 5,000 fpm (56.7 mph). Once the paper web is formed in a sheet,
its moisture content is reduced first using suction units, called the wet press
area, and later by drum type dryers. Paper sheet continuously runs through a
series of stainless steel drums heated up to 200oF (93oC) to ensure the sheet
has an approximate 4-5% moisture content.Depending on the type of paper,
further finishing processes are needed. Application of coats of various types of
chemicals applied to the surface of the paper make it extra shiny for special
applications, such as art papers. In general, coated papers are classified into
three groups: matte, semi matte and glossy.Finally, the paper sheet is wound
into large rolls, and then, they become ready to be shipped
Examination (physical, microscopic and chemical)