Lecture 7: Bodily fluids Flashcards
Bodily fluids
- Liquids ex/secreted by or present within the body at any given time
-Excreted –> e.g. faeces,
vomit, urine
-Secreted –> e.g.
blood/plasma, semen,
saliva
Forensic applications
- Within the human body
- Transferred to victim/scene
- Help forensic scientists determine:
-How/When a victim died
-Who the offender was
Forensic pathology
- Cause, manner, and time since death
-E.g. diatoms in the pleural
liquid
-E.g. stomach
contents/vomit analysis
-E.g. estimating time since
death using vitreous
humour
Fluids as evidence
- ‘Every contact leaves a trace’
- Direct vs indirect transfers
- Often have to assume presence
Forensic value
- Present at various crime scenes
- Persistence overtime
- Detection + identification= DNA analysis
-Individualisation
Detection
> Many fluids and stains are invisible
Screening techniques at scene
-E.g. alternative light
sources (ALS)
-E.g. chemical agents
(luminol)
Locate fluids ‘in situ’ before collection
Forensic approach
- Presumptive tests: indicate potential source
- Confirmatory tests: conclusively identify type of biological material
- Downstream analysis: individualise (DNA)
* Tests:
-Vary by type of fluid
-Non-destructive
-Scene-based/lab based
-Avoid contamination
Collection
> Scene, evidence, and/or persons
-Documentation
-Sensitive techniques-
swabs, tapes, cuttings
Comparison samples (controls)
Measures to avoid cross-contamination
Blood
> Most common bodily fluid encountered
-Murder, assault, burglary,
sexual offences, hit and run
Volume recovered varies- drop stain, pattern
Detectable after cleaning attempts
Various presumptive tests available
Uses of blood
- Forensic reconstruction
-Who was involved?
-What happened?
-When did it happen?
-What evidence is needed
from suspects?
Nature of blood
> Viscous- 3-4x thicker than water
55% plasma
-Water, antibodies,
proteins, enzymes,
hormones, amino acids,
glucose ….
-Inorganic substances=
drugs/alcohol
45% cellular material
Function of blood
> Erythrocytes
Leucocytes
Thrombocytes
Erythrocytes
> RBCs –> Red Blood Cells
Most common (44%)
Contain haemoglobin
-Iron containing protein
carriers O2 and CO2
-Gives blood its colour
Shape= flexibility for travel
Antigens on surface of RBCs
-Blood-type characteristics
Leucocytes
> WBCs –> White Blood Cells
4% blood volume
Contain nuclei= DNA
Protect against infectious disease + invaders
Five types of WBC
-Granulocytes (neutrophils,
eosinophils, basophils)
-Lymphocytes
-Monocytes
Thrombocytes
> Platelets
Cell fragments
Irregularly-shaped, colourless- produced in bone marrow
Sticky surface- forms clots –> stops bleeding
Fluidity of the blood
Active when circulatory system walls damaged
Blood types
> Classification of blood based on presence/absence of:
-antibodies (plasma)
-Inherited antigenic
substances (RBCs)
Blood type inherited from both parents
Two important systems:
-ABO
-RhD antigen
ABO systems
> Denotes presence of one, both, or neither A or B antigens on RBCs
Initially discovered by Landsteiner (1901)
Four main groups- A, B, O, AB
Phenotypes vary in different populations
Rhesus (Rh) system
> 2nd most important blood group system
50 defined blood group antigens incl. D, C, c, E and e
Initially discovered by Landsteiner and Weiner (1930’s) in monkeys
RhD antigen- important and most immunogenic
-Present (+), absent (-)
84% of Europeans RhD+ (ie A+, B+, O+, AB+)
AB(O)H secretors
> Person secrets antigens into bodily fluids
-Saliva, urine, tears, bile,
digestive fluids etc
80% of Caucasian population carry gene
Forensic importance:
-Blood group established
from other fluids
-Exclusion of non-secretor
-Useful history
Forensic significance of blood
> RBCs:
-Identify blood group
antigens
-Peroxidase-like activity=
detection
WBCs:
-DNA analysis
Plasma:
-Serum (species testing)
-Drug/alcohol screening
-BPA
Blood pattern analysis (BPA)
> Blood exits in the body as a liquid
Exits differently depending on injury/action
Analysis of patterns- what happened? Order of events?
Violent crime- Often copious amounts of blood
- Active bloodstain
> Blood travels by a force other than gravity
-Impact to body (weapon)-
spatter
-Projection (punctured
artery)- gushes or spurts
-Secondary object
(weapon)- cast off stains
Direction of travel on impact
-Tails
Angle of impact
-Round Vs elongated
- Passive Bloodstain
Blood formed solely under influence of gravity
> Blood flows often extensive
> Drops and pods
> Angle of impact
- Transfer Bloodstains
Blood deposit= direct contact with contaminated objects
> From weapon or person
> Disposing of evidence
> Indicate repeated contacts
Presumptive tests for blood
- Determine presence
-Trace levels
-Following clean up
-Test unknown stains
-At crime scene and in lab
1. Screening techniques
2. Catalytic tests
- Screening techniques for blood
> ALS/chemiluminescence (non-visible) traces
-E.g. Polilight, luminol,
fluorescein, Bluestar
Informs use of additional
presumptive tests
Rapid + non-destructive
ALS used to detect variety of fluids
- Catalytic tests for blood
> Haemoglobin catalyses oxidation of reagents (e.g. H2O2)
-Changes colour if +ive
Phenolphthalein (Kastle-Meyer tests) and leuchomalachile green (LMC)
Confirmatory tests for blood
> Absolutely identify unknown stain as blood
Microscopic assessment of RBCs and WBCs
Crytal tests- e.g. Teichman and Takayama
Spectroscopy highly reliable
Serological tests for blood
> Preciptin tests= identify human proteins
Further forensic analysis on blood type
Associate/exclude individuals
Saliva
> Common in burglary, assault, sexual offence cases
-E.g. bite-marks
-E.g. transferred to exhibits
(balaclavas, cigarettes etc.)
Non-invasive control sample collection
Forensic analysis:
-Identify the accused
-Detect drugs/alcohol
-Poisoning cases
-Hormone levels
-Species identification
(animal bites)
Nature of saliva
> Digestive acid- breaks down food starches
Produced by 3 main glands (sublingual, submandibular, partoid) –> serous and mucus cells
Humans produce 1-1.5L/day
99% water with pH of 6.8-7.0
-1% electrolytes,
immunoglobulins,
proteins, enzymes,
nitrogenous products,
WBCs ….
»_space;including digestive
enzyme: alpha amylase
Function of saliva
> Lubrication and cleansing of the mouth
Aids digestion:
-Broken up food= bolus
-Solubilise dry food
-a-amylase= breaks down
starch
-Salivary lipase=fat
digestion
Anti-microbial properties- e.g. lysozyme
Maintains oral hygiene
Forensic analysis of saliva
> Few well known and accepted presumptive tests
-Enzymatic methods
-ALS screening
No currently used confirmatory tests specific to saliva
Enzyme analysis of saliva
> Saliva ID based on presence of amylase enzyme
-Phadebas, SALIgAE, RSID
Sample of stain added to soluble starch solution, iodine added as regent
Not specific for human saliva
High false +ive rate
Semen
> Evidence in sexual offence cases
-Person (clothing, swabs,
combings, hair, blood)
-Scene (e.g. used condoms,
bedding, furniture)
Speed important- medical exam of victim
-Detectable up to 31 hr
(mouth), 44 hr (anus), 10
days (vagina), 19 days
(cervix)
Aim: identify/exclude a suspect
Nature of semen
> Complex fluid and cellular mixture produced by male reproductive glands
1. Cellular mixture= spermatozoa
-Reproductive cells carry
genetic material for sexual
reproduction
2. Complex fluid= seminal fluid
-Protective, nutrient rich
environment for sperm cell
post-ejaculation
15-30% from the prostate: proteins incl. acid phosphase, prostate-specific antigens & albumin
5% from epididymis & testis: sperm rich fluid
60-75% from seminal vesicles: hormones, proteins (coagulants, fructose, glands)
Function of semen
> Sexual reproduction in humans/animals
Contributes half of genetic information to offspring
Average ejaculate 2-6ml
-100-150 million sperm
cells/ml
Alkaline pH 7.2-7.6
Forensic analysis of semen
- Presumptive tests
> ALS- but not all semen fluoresces
> Tests for seminal acid phosphate (SAP) most common
-If present, AP will react
with alpha-naphthyl
phosphate
> Detect PSA (prostate specific antigens) e.g. p30
-Present in seminal
plasma= antibody-antigen
reactions - Confirmatory tests
> Microscopic ID of sperm cells
-Treated with stain to
visualise heads
-Christmas tree stain:
»_space;Nuclear Fast Red- nuclei
»_space;Picroindigocarmine-
tails
> RS-ID semen strip test
-Sensitive & specific
-Seminal vesicle-specific
antigen- semenogelin
-Pre-ejaculation fluid also
Vaginal fluid
> Not as common at crime scenes as blood, semen, saliva
Important role in sexual assault cases
Very few tests as fluid is not well defined
-Composition varies due to
menstrual cycle
-Variable hormone levels
Nature and function of vaginal fluid
> Vaginal secretions = mucus or fluid produced from cervical & vaginal glands
Average production- 6g/day
Protects from microbial/bacterial infections
Proteins incl. lysozyme, acid phosphase, amylase
Shed blood cells, urea, epithelial cells…
Presumptive tests for vaginal fluid
> No presumptive/confirmatory test routinely used
Detect glycogenated epithelial cells with periodic Acid-Schiff (PAS) reagent
-Stains cytoplasm magenta
-Reliability? Destructive?
-Rogers et al (2020)
Lactic/citric acid ratio- VF or semen?
Menstrual fluid
> Need to distinguish between peripheral & menstrual blood
Importance of context within the case- trauma? Natural bleeding?
Challenging distinction to make- new techniques developing (2017+)
SERATEC-PMB- detect haemoglobin and d-dimer
Quick, easy to use, preserves DNA
Forensic validation- assess effectiveness:
1. specific and sensitive (3ul)
2. Different treatment time based on age of sample
3. No false +ves or -ves (ages, mixtures)
4. Post-mortem +ve samples
5. DNA extracted- sample buffer and sample well
6. Successfully applied to case work samples
Urine
> Useful in (sexual) assault, harassment, drug, mischief, abuse cases
Difficult to detect
-Low sensitivity of available
tests
-False positive results
Less viscous than other fluids- dispersal, dilution
Odour not localised
Nature and function of urine
> Liquid by-product of the metabolism
Expels nitrogen rich by-products: urea, uric acis, creatinine
Average production: 1.4L/day
91-96% water
-Inorganic salts & organic
compounds (e.g. proteins,
hormones, metabolites)
-solids (e.g. urea)
Forensic tests for urine
> Diluted urine- limited ALS detection
Urea: activity of the enzyme urease
-Breaks down urea,
releases ammonia + CO2
-Detect ammonia via
Nessler’s or DMAC reagent
Creatinine: high concentrations
-Jaffe test= creatinine reacts
with picric in alkaline
medium
Limited efficiency with dried stains..
RSID urine test
> Tamm-Horsfall (THP) most abundant protein- 50-100mg/day excreted
Specific to urine but concentration varies
-Test= assay uses THP-
specific antibodies
-In-situ sampling increases
test sensitivity
-Blood presence inhibits
urine detection
-Not human specific
Sweat
> Less common bodily fluid found at crime scenes
Useful evidence in range of forensic cases
DNA evidence- extracted from hats, clothing, bedding etc,
Nature and function of sweat
> Fluids secreted by sweat glands in skin
Means of thermoregulation
Sweat rates up to 2-4L/hr or 10-14L/day
Mostly water with dissolved mineral traces
-Lactic acid
-Urea, creatinine
-NA, K, Ca, Mg
Forensic tests for sweat
> Most difficult of bodily fluids to identify
1. SEM-EDX: chlorine peak
basis of comparison
2. G-81 monoclonal
antibody (sweat-specific)
ELISA analysis
3. Metabolite biometrics:
single-analyte enzymatic
assays distinguish persons
Additional forensic intelligence- drug use, alcohol consumption
Forensic value of bodily fluids
> Fluids important for context of case- source
Downstream DNA analysis- enables robust identification
Fast, efficient, inexpensive- saves time, money, effort
Allows sorting of evidence by relevance
Body fluids analysis a major component of crime scene and lab processing
Proteomics
> Protein analysis= advances within bodily fluid testing
Proteome: proteins coded in genetic makeup of individual (genome)
-Study of all proteins in a
cell (structure,
modifications, interactions
etc.)
May yield detailed profile of an unknown individual
