This document discusses various body fluids and the presumptive and confirmatory tests used to identify them at a crime scene. It outlines tests for blood, saliva, urine, semen, fecal stains, and sweat. For blood, tests detect hemoglobin or heme compounds, including the luminol, fluorescein, phenolphthalein, Teichmann, and Takayama tests. Saliva tests identify the enzyme amylase, including using Phadebas tablets. Urine tests detect creatinine or urea. Semen tests identify acid phosphatase or prostate antigens. Fecal tests detect bilirubin breakdown products. And sweat can be identified using crystal violet dye interaction.

1. Body Fluids
Dr. Sangeetha Balakrishnan
23 February, 2017

2. Presumptive/Preliminary/Screening/
Field/Indicative Tests for Blood
1. Luminol Test
2. Fluorescein Test
3. Phenolpthalein Test

3. Luminol Test at a Crime Scene
0 to 30 s

4. Fluorescein Test at a Crime Scene

5. Phenolpthalein Test
• Can detect blood diluted to 1 part in 10
million.
• Gives false positives for certain vegetable
extracts.
• How is it done?:
(i) Suspected blood stain (on a cotton swab) +
water + Phenolpthalein colourless
(ii) Add hydrogen peroxide immediate
PINK colour!

6. Confirmatory Tests for Blood
1. Teichmann Test
2. Takayama Test (Hemochromogen Test)
• Both are called Microcrystal Tests.
Blood + Crystallising Reagent
Characteristic Shaped
Crystals.

7. Teichmann Test
• Blood + Teichmann’s Reagent (heat) Hb
hemin react with halides brownish-
yellow rhomboid crystals
• Microscopic Observation
• Teichmann’s Reagent: Potassium bromide +
Potassium chloride + Potassium iodide in
Acetic acid.

8. Teichmann Test

9. Takayama Test
(Hemochromogen Test)
• Suspected blood stain on a glass slide + heat +
add pyridine in Sodium hydroxide + reducing
sugar red feathery crystals of pyridine
ferroprotoporphyrin
• Very sensitive test.
• Even very old blood stains give +’ve results!

11. Saliva
• Saliva is produced in the mouth.
• Function: preliminary digestion of food.
• Contains: water, proteins, enzymes, salts.
• Alpha amylase: enzyme in saliva.
• Alpha amylase breaks down starch in food.
• It is also present in other body fluids, but in
low concentrations.

12. Preliminary Test - Saliva
• Principle: Starch + Iodine Deep blue colour
• Suspected saliva sample + water/saline
incubate at body temperature.
• If the suspected sample is indeed saliva, it will
contain alpha amylase!
• Alpha amylase will break down starch into
simpler components. (digestion)
• Add Iodine solution.
• Absence of starch (it has been broken down)
No Deep Blue colour!

14. Drawbacks of the Saliva
Preliminary Test
• Not particularly sensitive.
• Not specific to saliva.
• Use saliva sample for DNA testing.
• Detection of saliva at scene of crime:
Shine UV light fluorescence!

15. Saliva – Confirmatory test
• Phadebas Amylase Test
Developed by Pharmacia Diagnostics.
Qualitative and quantitative test.
Phadebas: a synthetic biochemical substrate.
• The substrate has starch microspheres.
• The microspheres are chemically bonded to a blue
coloured dye.
Phadebas substrate + suspected saliva (in water)
salivary amylase digests starch starch microspheres
break down blue dye is released!

16. Phadebas Amylase Test

17. Urine
• Main components:
(i) Creatinine
(ii) Urea
Presumptive Test – Urine
1. Jaffe Test
• Based on the detection of creatinine
• Suspected urine sample + picric acid + 5% NaOH
immediate orange colour

18. Presumptive Test – Urine … cont’d
2) Assay for Urea
• Bromothymol Blue is a pH-indicator dye.
• Colour: Yellow-green at pH 6
Aqua blue at pH > 7.6
(i) Whatman Filter Paper + Bromothymol Blue (1
drop) allow to dry
(ii) Add a drop from the suspected urine sample.
(iii) Add a drop of Urease.
(iv) If urea is present (meaning: urine is present),
urease will degrade urea to ammonia gas.
(v) This will cause the pH to increase.
(vi) Aqua blue colour!

19. Semen
• Produced by the male sex organ.
• Semen = Spermatozoa + Seminal fluid
(cellular component) (fluid component)
Spermatozoa
(Sperm)
Seminal Fluid
Male reproductive cells Contains: proteins, salts,
organic materials.
Flavin: UV fluorescence!
Consist of: head, tail and mid
piece
Produced in prostate gland
Seminal Acid Phosphatase
(SAP)
Prostate Specific Antigen (p30
protein)

20. Presumptive Test - Semen
• Based on Seminal Acid Phosphatase (SAP).
• There are other acid phosphatases in the
body!
• Brentamine Spot Test or Walker Test
• Suspected Semen Sample + Brentamine Fast
Blue B Intense purple colour within 2
minutes

21. Brentamine Fast Blue B added to
suspected semen samples

22. Confirmatory Test - Semen
1. Christmas Tree Stain Test
• Basis: Sperms analysed in lab are NOT motile.
• Hence identification should be in the presence of other
cellular material!
• A pair of dyes is used:
(i) Picroindigocarmine (PIC)
(ii) Nuclear Fast Red
• PIC: stains the tails green-blue-grey
• Nuclear Fast Red: stains the heads (with DNA) bright
crimson.

23. Christmas Tree Stain Test

24. Confirmatory Test – Semen … cont’d
2. Detection of Prostate Specific Antigen or p30
protein:
Oligospermia (low sperm count)
Aspemia (no sperms)
Suspected semen sample + Reagent Intense
purple.

25. Fecal Stains
• Basis: Human waste has bilirubin.
• Bacteria in body, break down bilirubin to
urobilinogen.
Presumptive Test: Edelman’s Reagent
Fecal stain bilirubin urobilinogen
urobilin shine UV light + Edelman’s reagent
Green fluoresence.

26. Confirmatory Test – Fecal Stains
• Microscopy:
• To look for animal and plant cells present due
to digestion.

27. Sweat
• Suspected sweat sample + Crystal Violet -
deep purple
• Fatty acids in sweat react with the dye.