Introduction to Microscopy

In this guide, we will explore the fascinating world of microscopy. We will cover the basics of light waves, the
properties of electromagnetic radiation, and the interaction of light with matter. We will also delve into the
use of refractive index in forensic science and the workings of lenses that help us see the tiniest of specimens.
By the end of this guide, you will have a better understanding of microscopy and its applications in a wide
range of fields.

ER by Erin Ross
The Parameters of Light Waves
Light waves have three measurable parameters: wavelength, amplitude, and frequency. The visible range of
light is from 400 to 700 nanometers, and different wavelengths appear as different colors. Amplitude
determines brightness intensity, while frequency is related to the energy of the wave.

We measure wavelength in Angstroms (Å), amplitude in candela (cd), and frequency in hertz (Hz). Simple
wave equations like c = f λ and E = hf help us calculate these parameters.
Forms of Electromagnetic Radiation
In addition to visible light, there are other forms of electromagnetic radiation, such as ultraviolet light and
infrared light. Unlike with visible light, these waves are transmitted as pure waves, not packets of energy
known as photons. Electromagnetic radiation can exist as longitudinal waves, which oscillate in the direction
of propagation, or transverse waves, which oscillate perpendicular to the direction of propagation
Refractive Index
Refractive index is a critical optical property used to determine the type of glass in forensic science. By
comparing RI values of glass fragments recovered from clothing to control samples, forensic scientists can
identify the type of glass involved in a crime. Refractive index is defined as the ratio of the speed of light in a
vacuum to the speed of light in a medium. We can calculate refractive index using the equation n = c/v, where
n, c, and v are the refractive index, speed of light in a vacuum, and speed of light in the medium, respectively.
Light and Matter Interaction
When light waves encounter matter, there are four ways in which they can interact: reflection, refraction,
absorption, and diffraction. Refraction is particularly important in microscopy, and the angle of incidence (i)
of an incident ray is related to the angle of refraction (r) of the refracted ray using the formula n1sin(i) =
n2sin(r), where n1 and n2 are the refractive indices of the media of incidence and refraction, respectively.
Lenses and Magnification
Lenses are spherical devices that refract light and can cause light to converge or diverge. Bi-convex lenses are
converging while bi-concave lenses are diverging. Magnifying glasses use convex lenses, which can produce a
magnified virtual or real image depending on the distance of the object from the lens. Magnification, the
degree by which the image dimensions are larger than the object dimensions, can be calculated using the
equation m=-di/do, where di is the image distance and do is the object distance.
Longitudinal Waves vs. Transverse Waves
Waves can be described as either longitudinal or transverse. Longitudinal waves oscillate in the direction of
propagation, such as sound waves, while transverse waves oscillate perpendicular to the direction of
propagation, such as light waves.
Speed of Light in a Vacuum
The speed of light in a vacuum is a fundamental constant that is approximately 299,792,458 meters per
second. This speed is a maximum and is usually denoted by the letter c. The speed of light in air is very similar
to the speed of light in a vacuum, and the refractive index of air can usually be assumed to be 1.
Simple Wave Equations
Two simple wave equations are used to calculate parameters of light waves. The equation c = fλ is used to
determine the speed of light in a vacuum (c), wavelength (λ), and frequency (f ), while the equation E = hf is
used to determine energy (E), frequency (f ), and Planck's constant (h). Calculating wavelength from
frequency using the first equation is essential for assuming the properties of electromagnetic radiation,
including radio waves, microwaves, ultraviolet light, and X-rays..
Coaxial and Divergent Light
Light sources can either be coaxial, where they have coaxial paths of propagation and are collimated to
minimize light scattering, or divergent, where they emerge unhindered from the source without any optical
instrumentation. To prevent divergence, an optical system is necessary.
The Law of Refraction
The Law of Refraction relates the angle of incidence (i) with the angle of refraction (r) of a refracted wave. It
states that n1sin(i) = n2sin(r), where n1and n2 are the refractive indices of the media of incidence and
refraction, respectively. By convention, positive values of power in lenses correspond to real images, objects,
and focal points, while negative values correspond to virtual objects, images, and focal points.
Speed of Light in a Medium
The speed of light in a medium is the speed at which light waves travel in that medium. We can define the
speed of light in a medium by using the ratio of the speed of light in a vacuum (c) and the refractive index (n)
of the medium, where v = c/n. By changing the refractive index of a medium, it is possible to change the speed
of light through that medium.
Microscopy Experimentation
Experiments in microscopy involve observing the interaction of light waves with matter. An understanding of
wave propagation, refraction, and reflection is essential for proper interpretation of microscopic images.
Careful sample preparation, proper lens selection, and the proper parameters (such as wavelength, speed of
light, and angle of incidence and refraction) are essential for obtaining accurate results.