Biomedical Engineering Reference
In-Depth Information
fashion. This facility obviates the necessity for the addition of tags, as is
often the case with fluorescence and electrochemical measurements.
Attachment chemistry at the device-liquid interface: The necessity for use
of gold as a substrate in SPR has meant that either dextran films or self-
assembly chemistry has dominated the methodology required to bind
biomolecules to the chip. AW is similar with respect to the gold electrodes
in TSM technology, but offers more flexibility with Si chemistry for
quartz. The dextran layer is regarded by some to be problematical in that
analyte diffusion is influenced by the polymer.
Sensing mechanism: For the AW case, transfer of energy to the liquid
implies an acoustic couple between the device surface (and imposed
chemistry) and the surrounding liquid. Accordingly, the measured
properties of AW sensors, such as resonant frequency, are governed by a
number of interfacial properties, such as surface-free energy, interfacial
viscosity and charge. Therefore, these devices are exquisitely sensitive to
the changes in interfacial chemistry associated with biochemical reactions
instigated at the sensor-liquid interface. These observations confer
possibilities to detect conformational shifts and the binding of very small
molecules. For SPR, the resonance condition is extremely sensitive to
variations in the refractive index immediately adjacent to the metal film,
which is both a blessing and a hindrance! There is a level of debate as to
the ability of SPR to detect small molecules, especially when they are
attaching to an already bound macromolecule.
Sensitivity and limit of detection: This is far from straightforward because
incorrect terminology and reference points are prevalent in the literature
and wide ranges of values are quoted. A monolayer of protein adsorbed to
an EMPAS surface yields a change in frequency of 5000Hz with noise in
frequency being around 100Hz. In terms of solution concentration the
limit of detection (LOD) would be in the region of 10
11
M. Given the
capability of SPR with respect to minimum value of RIU, the LOD
number is generally regarded to be 10
12
M.
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1.4.3.3 Interferometry
Interferometry is based on the superposition of, usually, electromagnetic waves,
which yields information concerning the original nature of the waves. The
reader will be familiar with the physics of the Michelson interferometer, which
is at the heart of the Fourier transform infrared spectrometer. (With light of
this wavelength, superposition can be examined through the use of a moving
mirror for one of the light pathways.) In biosensing technology several types of
device designs have been employed to attempt the detection of biochemical
species, examples being Mach-Zehnder, Young and Hartman sensors
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(depicted in Figure 1.28(a), (b) and (c), respectively).
The Mach-Zehnder apparatus involves two different light pathways, as is
typical in interferometry, with one being subjected to passage through the
ample. The laser radiation is then combined with a reference beam resulting in
