This document discusses methods for estimating the age of bloodstains, which can help determine the timing of criminal events. It describes using Raman spectroscopy to analyze bloodstain spectra and date stains that are less than two years old. An alternative method is fluorescence lifetime measurements, which takes advantage of changes in tryptophan fluorescence as blood proteins undergo structural changes during aging. Both Raman spectroscopy and fluorescence lifetime analysis provide accurate, non-destructive ways to estimate the age of a bloodstain.

2. AGE ESTIMATION FROM
BLOODSTAIN
- DIKSHA MEHRA
MEMBER AFRS

3. CONTENTS
• Blood
• Blood Stain Detection
• Different Bloodstains
• Age Estimation
• RAMAN Spectroscopy
• Dating with Flourescence
• References
Fig. 1 Bloodstain

4. Blood
• A bodily fluid. Connective
tissue made up of different
types of cells carrying
nutrition and blood gases.
Fig.2 Blood Composition

5. BLOODSTAIN DETECTION
Preliminary Test For Blood
• TMB or Benzidine Test
• Phenolphthalein Test
• Luminol test
Fig.4 Blood Preserving
Tubes
Fig.5 FTA Card

6. Different Bloodstains
Fig.6
Different
Types of
Bloodstains

7. AGE ESTIMATION
• Bloodstains have a significant role to play as these can determine
the exact duration from the time of occurrence till the found
time.
• Methods :
- RAMAN Spectroscopy
- Dating Bloodstains with Fluorescence Lifetime
measurements

8. AGE ESTIMATION – RAMAN SPECTROSCOPY
• Raman spectroscopy and
advanced statistics allow to date
a blood stain accurately, provided
said blood stain is less than two
years old.
• Raman spectroscopy involves
shining a laser on a sample and
measuring the intensity of
scattered light.
• No two substances give the exact
same Raman spectrum and so the
spectrum is essentially a unique
fingerprint for a given material.
• Importantly, from the criminal
evidence point of view, the
process is non-destructive.
• Thus, it can be repeated at any
time, and also allows other tests,
such as DNA sequencing, to be
carried out.

9. DATING BLOODSTAINS WITH FLUORESCENCE
LIFETIME MEASUREMENTS
• The method is fast, reproducible,
concentration independent, and
requires a miniscule amount of
the bloodstain with minimal
sample preparation.
• The approach is based on the fact
that the fluorescence lifetime of
tryptophan the major endogenous
fluorophore in blood proteins is
highly sensitive to the protein
conformation.
• In the process of blood aging, the
major tryptophan-containing
proteins, such as albumin and γ-
globulins, undergo structural
changes.
• Therefore, the time-related
degradation of proteins in blood
reflects the blood age.
• This approach minimizes
problems with sampling and
provides more accurate results.

10. REFERENCES
• https://pubs.acs.org/doi/10.1021/acs.analchem.8b01367#:~:text
=In%20addition%2C%20the%20age%20of,analyzing%20the%20color%2
0of%20bloodstain.&text=There%20is%20a%20technique%20to,%2C24
)%20and%20decomposition%20degree.
• https://www.chemistryviews.org/details/ezine/1435459/CSI_The_
Age_of_the_Bloodstain.html
• https://chemistry-
europe.onlinelibrary.wiley.com/doi/abs/10.1002/chem.201102935