These results themselves do not furnish us with much information
beyond a confirmation that our theoretical model for energy transfer
between the dyes is valid. However, Lehn (in Supramolecular Chemistry,
VCH Weinheim, 1995) presents a compelling application for such
behavior in dye assemblies. Consider two antibodies that both bind
specifically and at a different site to an antigen. By coupling dyes A and D
to the two different antibodies, the dyes become sufficiently close to allow
energy transfer. By introducing an excess of the modified dyes, spectroscopic analysis could be used to perform a simple immunoassay. After
accounting for background fluorescence from modified antibodies
unbound to the antigen interacting with one another directly, fluorescence from acceptor A could be monitored and would indicate the specific concentration of the antigen in question.
9.3.2 Photorelaxation
The functionality of the aforementioned dye assemblies hinges on the
differences in absorbance and emission spectra, or Stokes shift, of the two
entities. We have already discussed the Stokes shift in the context of
Raman spectroscopy; it is a shift in energy of a photon due to a transition
between vibrational levels when a molecule is excited to, relaxes from, or
scatters photons via an excited electronic state. As we discussed in
Chapter 6, when a molecule is excited to a higher electronic state, it can
relax via fluorescence, but often undergoes nonradiative vibrational
relaxation first, losing energy to its environment. The typical lifetimes of
excited electronic states are very short, often on the order of 10 ns, after
which the molecule reverts to its ground state. When the molecule relaxes
to the ground state, it emits energy corresponding the energy difference
between the excited state and ground state in the form of a photon of the
corresponding wavelength, as per Einstein’s relation
E photon =
hc
l
(9.9)
where h = 6.62606896 × 10
−34 J s and c = 2.998 × 10
8 m/s. The difference
between the excitation photon wavelength necessary to induce an electronically excited state and the wavelength of the subsequently emitted
photon is known as the Stokes shift.
ΔE stokes = hc
1
l excitation
−
1
l emission
(9.10)
EXCITATION TRANSFER 351
beyond a confirmation that our theoretical model for energy transfer
between the dyes is valid. However, Lehn (in Supramolecular Chemistry,
VCH Weinheim, 1995) presents a compelling application for such
behavior in dye assemblies. Consider two antibodies that both bind
specifically and at a different site to an antigen. By coupling dyes A and D
to the two different antibodies, the dyes become sufficiently close to allow
energy transfer. By introducing an excess of the modified dyes, spectroscopic analysis could be used to perform a simple immunoassay. After
accounting for background fluorescence from modified antibodies
unbound to the antigen interacting with one another directly, fluorescence from acceptor A could be monitored and would indicate the specific concentration of the antigen in question.
9.3.2 Photorelaxation
The functionality of the aforementioned dye assemblies hinges on the
differences in absorbance and emission spectra, or Stokes shift, of the two
entities. We have already discussed the Stokes shift in the context of
Raman spectroscopy; it is a shift in energy of a photon due to a transition
between vibrational levels when a molecule is excited to, relaxes from, or
scatters photons via an excited electronic state. As we discussed in
Chapter 6, when a molecule is excited to a higher electronic state, it can
relax via fluorescence, but often undergoes nonradiative vibrational
relaxation first, losing energy to its environment. The typical lifetimes of
excited electronic states are very short, often on the order of 10 ns, after
which the molecule reverts to its ground state. When the molecule relaxes
to the ground state, it emits energy corresponding the energy difference
between the excited state and ground state in the form of a photon of the
corresponding wavelength, as per Einstein’s relation
E photon =
hc
l
(9.9)
where h = 6.62606896 × 10
−34 J s and c = 2.998 × 10
8 m/s. The difference
between the excitation photon wavelength necessary to induce an electronically excited state and the wavelength of the subsequently emitted
photon is known as the Stokes shift.
ΔE stokes = hc
1
l excitation
−
1
l emission
(9.10)
EXCITATION TRANSFER 351