My research revolves around fluorescence spectroscopy. So what
is fluorescence spectroscopy and why is it useful?
What is fluorescence?
We are well aware of the fact that light interacts with matter.
Light interacts with cola in a different way to, say, carbonated
spring water. Visible electromagnetic radiation is detected by the
rods and cones in our retinas so that we can appreciate the difference
between water and cola. Plants absorb the sun's radiation in the
process known as photosynthesis - this is a different way in which
light interacts with matter but the important word in describing
these interactions is "Absorb".
The story of how matter interacts with light does not end with
absorbance. A small set of materials are also able to fluoresce.
This means that after taking light in (the absorbing process) they
are able to emit the light a small fraction of a second later; they
send back out light of a different color.
What are some good examples of fluorescence? Well actually there
are very few examples to speak of. However, a good analogy of what
is taking place in the emission process is the charging and discharging
of glowstars. When I was a boy I had a collection of glow-stars
on my bedroom ceiling. I would charge up my glowstars (allow my
glowstars to absorb) by exposing them to my bedroom lamp. When I
switched off the light the stars would glow; I could then watch
them and eventually fall asleep. Fluorescence spectroscopy allows
me to measure the colors of the light that are given off by a sample
that has already been charged by absorption.
A diagram of the fluorescence spectroscope or fluorometer is given
below. Each "monochromator" is simply a device used to
select a specific color or "wavelength of light".
Fluorescence efficiency (Quantum yield) is an intrinsic property
of a material. Hence we can monitor the presence of a given "fluorophore"
(something that fluoresces). Furthermore, fluorescence efficiency
and the fluorescence spectrum may often change depending on the
fluorophore's environment, or the material's conformation or morphology.
Here are some specific examples.
A set of polymers having a conjugated backbone (alternating single
and double bonds) are able to both conduct electricity and to fluoresce.
These polymers have found commercial application in organic light
emitting diodes that can be ink-jet printed onto flexible conductive
substrates. The efficiency of these devices has been improved by
studying the fluorescence of the polymer. The fluorescence efficiency
has improved as a result of chemical modification and fluorescence
measurement but, as well, the alignment of the polymer chains has
been investigated through use of spectroscopy. The alignment of
these chains, resulting in increased order has led to improved conductivity
in such devices. These conjugated polymers are also being used today
as a host matrix for photovoltaics (solar cells).
Tryptophan is a naturally occurring amino acid that fluoresces.
If the tryptophan amino acid is buried in a hydrophobic (oily) portion
of a protein its fluorescence will be different when compared to
a tryptophan that is exposed to the surrounding solvent, water.
Hence, at a simple level one can obtain information about the structure
of a protein. More importantly the tryptophan fluorescence may be
quenched by the presence of a substrate that the protein is acting
upon. The intensity of the fluorescence can be used to understand
how the protein is functioning, and how its conformation is changing
as it functions.
Many fluorescence tags are available that are selective in their
ability to bond to substances of (biological) importance. In flow
cytometry, for example, fluorescent tags stick only to diseased
cells inside a blood sample. By measuring the amount of fluorescence
the number of diseased cells in a patient's sample can be determined.