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

2. Presumptive/Preliminary/Screening/ Field/Indicative Tests for Blood
Luminol Test
Fluorescein Test
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
Whatman Filter Paper + Bromothymol Blue (1 drop) allow to dry
Add a drop from the suspected urine sample.
Add a drop of Urease.
If urea is present (meaning: urine is present), urease will degrade urea to ammonia gas.
This will cause the pH to increase.
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
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:
Picroindigocarmine (PIC)
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.

28. Blood Spatter Analysis

29. Blood Stains
Passive Bloodstain
Eg. Clots
Drops
Flows
Pooling
Transfer Stain
Eg. Wipes
Swipes
Pattern Transfers
Projected or Impact
Stains
Eg. Spatters
Splashes
Cast-off stains
Arterial spurts
or gushes

30. arterial spurts
splash
passive fall
blood pool
multiple drips
smear

31. Physical Properties Viscosity – more viscous than water
Does not spatter – smooth hard surface (tile)
Spatters – rough, hard surface (concrete)
Surface tension – blood has a high surface tension; causes a decrease in its surface area.
Blood adheres to an external surface; it will separate and spatter only when there is sufficient external force to overcome
the surface tension.
Blood drops that fall on a flat surface have a spherical surface
Edges may have spikes or extensions
Satellites - small secondary droplets

32. Cast off pattern: blood from a moving object coated in blood (pipe, knife)
Fine-mist spatter: high-velocity impact (gunshot)
Void: empty space; victim/attacker/object moved after attack

33. Blood Spatter Types High velocity – gunshot wounds
Medium velocity – beating, stabbing
Low velocity – blunt object impact

34. Information from Blood Spatter
Used to explain events at a violent crime scene
Point of origin
Angle of impact
Blood velocity (manner of death)

35. Lines of Convergence (Point of Origin)
Determine source of blood (point of origin)
Draw straight lines down long axis of blood spatter

36. Angle of Impact
Determined by measuring the length and width of the blood stain.
arcsin A = Width/length
Here A = Angle of impact.