This document discusses the analysis and examination of blood stains from a forensic perspective. It outlines various physical, chemical, and microscopic tests that can be used to analyze blood stains, including benzidine tests to detect the presence of blood, microscopic examination to look for red blood cells, and precipitin tests to determine blood type. It also discusses how blood stain analysis is important for medicolegal cases involving identification of victims or suspects, crime scene reconstruction, and questions of paternity or inheritance.

1. Blood stain
Dr. GURJEET SINGH
PGT
AMCH

2. Agenda
Physical Examination
Chemical examination
Microscopic examinatio
Precipitin Tests
ABO System
Medico-legal aspects
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3. Introduction
Forensic serology involves the
examination and analysis of a variety of
body fluids which includes blood, saliva,
semen and urine.
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4. COLLECTION
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A clean piece of white
filter paper, or piece of
white clean cloth or
gauge & allowed blood
to soak in it, then
drying.
If object is porous, a
portion of unstained
area should be taken.
If object is non-porous,
particularly if its
metallic stains can be
removed by scraping
and place on a small
glass containers.
Stains on the cloth,
scrapped off or a
fragment or a material
cut.
If blood is liquid a
sample can be pipetted
and place in a tube and
refrigerated.

5. Analysis of blood
Blood is a complex fluid with pH-
7.4 cells about 45% and plasma
about 55%.
Legal requirements that
identification of the stain should
be established to a scientific
certainty.
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7. Physical examination
The valuable clues can be achieved
by visiting crime scene.
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8. Clothing
and
stains on
it
Size,
shape &
direction
of blood
spots
Character
of stains
Age of
stains
Condition
of body
Other
causes of
stains
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Diagrammatic representation of types of blood spots
commonly encountered at a scene showing
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10. Chemical examination
• Benzidine Test (Adler) and phenolphthalein test (Kastle Meyer test) is used to find whether the stain is blood or
not. (enzyme peroxidase).
• When benzene solution is added to the suspected solution a greenish blue colour is obtained if blood is present.
- Benzene solution is a combination 10% benzidine in glacial acetic acid and hydrogen peroxide.
. Test can remain positive if 1:500,000 dilution of blood is made.
. Benzidine is carcinogenic in nature that’s why some labs preferred
other test.
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11. Phenolphthalein test (Kastle Meyer test)
• Kastle Meyer test colour tests are based on a similar principal to luminol which reacts with
hydrogen peroxide in the presence of blood.
• In this case the chemical involved Phenolphthalein. This will react with hydrogen peroxide in the
presence of haemoglobin to produce a pink colour.
H2O2 PINK COLOUR
HEME PHENOLPHTHALEIN
H2O O
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12. OTHER TESTS
O- Tolidine ( Kohn or
O’Kelly) test
Tetramethylbenzidine.
Leucomalachite
green.
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13. Microscopic Examination
Red Corpuscles: Intact red
cells are seen only when the
stains are fresh or when
blood clot is available.
The red is unrecognizable
when drieds
A small piece of stain is cut
and soaked in a watch glass
with 2 – 3 drops of vibert's
fluid (Sodium chloride 2g,
mercuric chloride 1/2g,
distilled water 100ml) or
normal saline for half an
hour then examined under
microscope.
If the stain is not dissolved,
dilute solution of ammonia
or 2% hydrochloric acid can
be used.
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14. 14
Hen Human
frog
Bird
Cat

15. Confirmatory Test
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16. Teichmann Test
Place a sample suspected blood on glass slide, add few crystals of sodium chloride and
a few drops of glacial acetic acid from the side of the cover slip and heat it form a
hematin derivative.
These Hemin and hematin chloride crystals are brownish rhombic shaped, arranged
singly or in clusters.
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17. Haemin Crystals

18. Place a small stain
sample under a
coverslip and allow
the Takayama
reagent (sodium
hydroxide, pyridine
and glucose) to flow
under and saturate
the sample.
After a brief heating
the crystals are
viewed.
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Pink Feathery Crystals

20. Spectroscopic Examination
It is the most delicate and
reliable test for detecting
the presence of blood in
both recent and old stains.
Less than 0.1 mg of blood
is sufficient.
Blood stain is dissolved in
water, normal saline or
dilute ammonia.
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22. Serological Examination
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1) Precipitin test :- blood serum contains proteins in
colloidal suspension, and when human serum is injected in
to the animal becomes immunised against these proteins and
antibodies develop in the blood

23. Latex test
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A Saline extract of bloodstain is mixed
with dilute suspension of latex particles
sensitized with antiserum. A positive
reaction is shown by agglutination of
the particles into clumps.

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Fine Line of precipitate at the
interface of the two solutions

25. Hemagglutination Test
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When Human globulin
is mixed with
antihuman globulin, the
latter is absorbed and is
no longer capable of
agglutinating Rh
positive red cells
sensitized with
incomplete anti-d. this
detects globulins.

26. Age of the stain
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Fresh stain – light coloured
cloth appears bright red, is
moist and sticky.
Turns reddidsh brown in 24
hours.
Darn brown or black –
more than 24 hours

27. Venous blood is dark
red.
Arterial blood is
bright red.
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28. Grouping of blood stains
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31. MEDICO –LEGAL ASPECTS
• Disputed Paternity:- The question of
disputed paternity arise in the court in
the following conditions:
• Adultery and divorce
• Blackmail
• Maintenance claims
• Share of property
Civil Cases
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33. Criminal Cases
Identification of
victim or
offenders.
Stains due to
body fluids : The
blood group
antigens can be
demonstrated in
stains on clothes
due to semen,
sweat or saliva
(secretors)
Crime scene
reconstruction
Malingering Cause of death Time since death
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34. Reference
1)Saukko P, Knight B. Knight’s forensic pathology.4th Ed. CRC
Press; 2015
2)DiMaio VJM, Kimberley Molina D. DiMaio’s Forensic
Pathology. Boca Raton: CRC Press; 2021.
3)Subrahmanyam BV. Parikh’s textbook of medical
jurisprudence, forensic medicine and toxicology for
classrooms and courtrooms. 8th ed. New Delhi, India: CBS
Publishers & Distributors; 2019.
4)Reddy KSN, Reddy M. The synopsis of forensic medicine &
toxicology the synopsis of forensic medicine & toxicology. New
Delhi, India: Jaypee Brothers Medical; 2022.
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35. Thank you