Ifugao State University

COLLEGE OF CRIMINAL JUSTICE EDUCATION

Potia Campus, Alfonso Lista Ifugao

INSTRUCTIONAL MATERIALS in FORENSIC INSTRUMENTATION

Part II
A. The polarizing microscope
 In 1808, French physicist Etienne Louis Malus discovered the refraction and
polarization of light.
 In 1828, William Nicol of Edinburgh invented a prism for polarization, as an
indispensable part of polarizing.
 In 1839, William Henry Fox Talbot invented polarizing microscope
 A compound or stereoscopic microscope can be modified to be outfitted
with a polarizer and analyzer so as to be capable of allowing the viewer to
detect polarized light.
 The effect of introducing a specimen that polarizes light will be to orient the
polarized light, allowing it to pass through the analyzer. The result produces
vivid colors and intensity contrast that make the specimen readily
distinguishable. Usually used in the field of geology.
 Produces 3D images
 Works on the principle of interference

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 Polarizer, positioned in the light path somewhere before the
specimen, and:
Analyzer (a second polarizer), placed in the optical pathway
between the objective rear aperture and the observation tubes or camera
port. Image contrast arises from the interaction of plane-polarized light with;
Birefringent (or doubly-refracting) specimen to produce two
individual wave components that are each polarized in mutually
perpendicular planes. The velocities of these components, which are
termed the ordinary and the extraordinary wavefronts, are different and
vary with the propagation direction through the specimen. After exiting the
specimen, the light components become out of phase, but are recombined
with constructive and destructive interference when they pass through the
analyzer.
B. The micro spectrophotometer
 First microspectrophotometer was built in the 1940s.
 By linking a microscope to a computerized spectrophotometer, a new
dimension has been added to the capability of the microscope, giving rise
to the micro spectrophotometer.
 Using the micro spectrophotometer, a forensic analyst can now view a
particle under a microscope while, at the same time, a beam of light is
directed at the particle in order to obtain its absorption spectrum.
 Designed to measure UV-Visible-NIR spectra of microscopic samples or
microscopic areas of larger objects. (Note: NIR= near infrared)
 To measure UV-visible-NIR range transmission, absorbance, reflectance,
emission and fluorescence spectra of sample, you need the CRAIC
software.
o This allows identification of
- Dyes
- Materials
- Concentration of chemicals

ELECTRON MICROSCOPE
In 1931, German scientist Ernst Ruska developed the first electron microscope.
The Electron Microscope (EM) is an impressively powerful microscope that exists today,
allowing researchers to view a specimen at nanometer size.
They utilize the same principles behind an optical microscope, but rather than photons or
particles of light, concentrate electrons, charged particles located on the outside of atoms,
onto an object.
Additional differences include preparation of specimens before being placed in the
vacuum chamber, the use of coiled electromagnets instead of glass lenses, the use of a
thermionic gun as an electron source and the image or electron micrograph is viewed on
a screen rather than an eyepiece.
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 Techniques, which vary based on type of specimen and analysis, include:

 Cryofixation - a technique for fixation or stabilization of biological

materials as the first step in specimen preparation for electron microscopy
and cryo-electron microscopy.
 Fixation - Fixation of tissues is the most crucial step in the preparation of tissue
for observation in the transmission electron microscope.
a. Osmium tetroxide and aldehydes
b. Sodium acetate/sodium veronal
 Dehydration – remove water from the biological specimens so that they
can be further processed using light microscopy and electron microscopy.
 Embedding – tissues to be analyzed in the electron microscope are
usually embedded in resins for ultramicrotomy.
 Epoxy resins
 Acrylic resins
 Sectioning - produce clear images of focal planes deep within a thick
sample.
 Staining - the most widely used stains in electron microscopy are
the heavy metals, uranium and lead.
 Freeze-fracture and Freeze-etch - a process whereby specimens,
typically biological or nanomaterial in nature, are frozen, fractured, and
replicated to generate a carbon/platinum “cast” intended for examination
by transmission electron microscopy
 Sputter Coating - is the process of applying an ultra-thin coating of
electrically-conducting metal – such as gold (Au), gold/palladium (Au/Pd),
platinum (Pt), silver (Ag), chromium (Cr) or iridium (Ir) onto a non-
conducting or poorly conducting specimen.
C. Transmission Electron Microscope (TEM)
Like the scanning electron microscope, the transmission electron
microscope (TEM) uses electrons in creating a magnified image, and samples
are scanned in a vacuum so they must be specially prepared. Unlike the SEM,
however, the TEM uses a slide preparation to obtain a 2-D view of specimens,
so it's more suited for viewing objects with some degree of transparency. A TEM
offers a high degree of both magnification and resolution, making it useful in the
physical and biological sciences, metallurgy, nanotechnology and forensic
analysis.
The TEM is a popular choice for nanotechnology as well as semiconductor
analysis and production.

D. The Scanning Electron Microscope (SEM) is a different and final approach to

microscopy that will be considered.
The image formed by the scanning electron microscope is produced by
targeting, with electromagnets, a beam of electrons onto the specimen and
studying the electron emission on a closed TV circuit. The primary electron beam,
emitted from a hot tungsten filament, causes the emission of electrons from the
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 element making up the outer layer of the specimen. The emitted electrons are
collected, and the integrated and amplified signal is displayed on the TV circuit.

- The major attractions of the SEM image are its high magnification, high
resolution, and great depth of focus.
- In its usual mode the SEM has a magnification range of 10X to 100,000X.
- Its depth of focus is some 300X better than optical systems at similar
magnifications, and the resultant picture is almost stereoscopic in appearance.
- Its great depth of field and magnification are invaluable in determining structural
relationships over a contextually broad area.
- Microscope examination of fibers, like the cotton fiber using SEM, cannot
provide quantitative characteristics of fibers, but is non-destructive and can be
used for a qualitative comparison.
Although SEMs are approximately 10 times less powerful than TEMs, they produce
high-resolution, sharp, black and white 3D images.

Electron Microscope Advantages

 The primary advantage is its powerful magnification.
 The potential runs the gamut of scientific fields including biology, gemology,
medical and forensic sciences, metallurgy and nanotechnologies.

 EMs also have many technological and industrial applications, such as

semiconductor inspection, computer chip manufacturing, quality control and can
even be used as part of a production line.

Electron Microscope Disadvantages

 The main disadvantages are cost, size, maintenance, researcher training and
image artifacts resulting from specimen preparation.

 This type of microscope is a large, cumbersome, expensive piece of equipment,

extremely sensitive to vibration and external magnetic fields.

 It needs to be kept in an area large enough to contain the microscope as well as

protect and avoid any unintended influence on the electrons.

 Upkeep involves maintaining stable voltage supplies, currents to electromagnetic

coils/lens and circulation of cool water so the samples are not damaged or
destroyed from heat given off during the process of energizing the electrons.

 Special training is required to learn the involved processes of specimen

preparation, to minimize and recognize preparation-related artifacts and to
operate the microscope itself.

 Despite these disadvantages, EMs are assets to high-end research laboratories;

this powerful piece of equipment has resulted in innumerable advances in science
and industry.

Compiled by: Jonalyn Otongngan