This presentation is about electrophoresis and its forensic application

2.  Electrophoresis is a physical method of analysis which
involves separation of the compounds that are capable
of acquiring electric charge in conducting electrodes.
 Electrophoresis is an important analytical tool used in
all branches of biology. Electrophoresis was observed
for the first time in 1807 by Russian professors Peter
Ivanovich Strakhov and Ferdinand Frederic Reuss
at Moscow State University, who noticed that the
application of a constant electric field
caused clay particles dispersed in water to migrate

3.  Electrophoresis may be defined as the migration of the
charged particles through a solution under the
influence of an external electrical field.
 Ions that are suspended between two electrodes tends to
travel towards the electrodes that bears opposite charge.
 THE AIM OF CARRYING OUT ELECTROPHORESIS
INCLUDE :
• To determine the number, amount and mobility of
components in a given sample or to separate them.
• To obtain information about the electrical double layers
surrounding the particles.
• Determination of molecular weight of proteins and DNA
sequencing.

4. PRINCIPLE OF ELECTROPHORESIs
 Comprehensive term that refers to the migration
of charged particles of any size in liquid medium
under the influence of an electric field.
 Depending on kind of charge the molecule carry,
they move towards either to cathode or to anode.
 The rate of migration of an ion in electrical field
depending on factors:
• Net charge of molecule.
• Size and shape of particle.
• Strength of electrical field.

5. • Properties of supporting medium
• Temperature of operation
 This can be represented by the following equation
V=E.q/f
 For molecules with similar confirmation , f varies
with size but not with the shape. The
electrophoretic mobility (µ) of a molecule is
directly proportional to the charge density
(charge/mass ratio).
 Molecules with different charge/mass ratio
migrate under the electric field at different rates
and hence get separated.

6. TYPES OF ELECTROPHORESIS
1) Zone Electrophoresis
 Paper Electrophoresis
 Gel Electrophoresis
a- AGE
b- PAGE
c- SDS PAGE
d- 2D Electrophoresis
 Thin layer Electrophoresis
 Cellulose acetate Electrophoresis
2) Moving boundary Electrophoresis
 Capillary Electrophoresis
 Isotachophoresis
 Immuno Electrophoresis
 Isoelectric Focusing

7.  In this the sample is applied as a circular support on a strip
of whatman filter paper moistened with the buffer solution.
 The ends of the paper are immersed in separate reservoirs
containing buffer and in which the electrodes are fitted.
 Upon passing electric current, the ions in the sample
migrate towards oppositely charged electrodes.
 This method is suitable for separation of low molecular
weight compounds such as amino acids and nucleotides.

8. Forensic APPLICATIONS OF PAPER
ELECTROPHORESIS
 Paper Electrophoresis is used to identify and
separate drugs in the field of FORENSIC
CHEMISTRY.
 This technique is also used to isolate and compare
hemoglobin in the field of FORENSIC
HEMATOLOGY.

9. • Separation is brought about through molecular sieving
technique, based on the molecular size of the substances.
Gel material act as a “molecular sieve”. Gel is a colloid in
a solid form (99% is water).
• Different types of gels which can be used are; Agar and
Agarose gel, starch, polyacrylamide gels.
• A porous gel acts as a sieve by retarding or, in some cases
by completely obstructing the movement of
macromolecule of while allowing smaller molecules to
migrate freely.

10. a. AGAROSE GEL ELECTROPHORESIS
 Agarose gels are more porous and have a large pore size and
therefore used to fractionate large macromolecules such as nucleic
acids.
 Agarose gels are cast by boiling agarose in presence of a buffer
then poured into a mold and allowed to harden to form a matrix.
 Higher the agarose larger is the pore size.
 When the electric feild is applied across the gel, negatively
charged molecules move towards oppositely charged electrodes on
the basis of their molecular size.
 Smaller DNA molecules travel faster than larger ones. Commonly
used buffers- Tris Acetate, Tris Borate or Tris Phosphate at a
concentration of 50mM over a pH range of

11. b. POLYACRYLAMIDE GEL ELECTROPHORESIS
 The components used in the formation of this gel are known to be
neurotoxins.
 The most commonly used components to synthesize the matrix are
acrylamide monomer, ammonium persulphate and
tetramethylenediamine (TEMED).
 These free radicals can activate acryl amide monomer including them to
react with other acrylamide monomer forming long chains.
 This chains become cross-linked if the reaction is carried out in the
presence of N-N’-methylenebisacrylamide.

12. Forensic APPLICATIONS OF GEL ELECTROPHORESIS
 Gel electrophoresis used in FORENSIC science is a way
to analyze DNA. Because the DNA of each person is
unique, the patterns of separation created using Gel
Electrophoresis are unique.
 In the case of a crime, the forensic scientist can perform
an Electrophoresis of DNA from the crime scene and
from the suspect's DNA, then determine if the two are an
exact match.

13. 3. CAPPILARY ELECTROPHORESIS
 The technique that has very high resolving power and
speed.
 CE involve Electrophoresis of sample in small narrow
capillary tubes and high voltage from kV is applied.
 This technique is used to separate biological material
including amino acids, proteins, nucleic acids as well as
drugs or even metals. Very small amount of sample are
required for analysis.

14. FORENSIC APPLICATION of CAPILLARY electrophoresis
 Used for FORENSIC drugs, toxicological samples, fibres
analysis.
 Used for the analysis of explosive and gunshot residues.

15. Case Study
 In FORENSIC Science many biological evidences play an
important role, as they are the source of DNA.
 The DNA is isolated from different sources which include Blood,
Blood stains, Semen, Tissues, Bones and Hair in different cases
like Rape, Missing identity, Murder and Paternity.
 In this sexual assault case study blood and semen are the source
of DNA for DNA Finger Printing.
 A girl aged 23 had been brutally raped by a person of age 26 at
Kurnool district in Andhra Pradesh, India.
 After filing a complaint by the girl, the semen stained under
garment wore by the girl at the time of rape, was collected, DNA
was isolated from the semen in the inner ware and the STR
profiles were taken as Electropherograms by using Capillary
Electrophoresis technique.
 On the other hand Blood sample was collected from the suspect
in this rape case, DNA was isolated from it and STR profiles were
taken as Electropherograms.
 Both the Electropherograms are compared and the suspect is
proved to be the culprit in this case.