Gilchrist and Reynolds
166
However, this type of approach is not realistic to the normal use of monochromators
as broad-band tunable light sources or broad-band detection system. Typically, stray light
is very difficult to measure, as it strongly depends on the wavelength, bandpass used, and
the type of source. A further discussion of this important point is made in Section 5.4.8 on
instrument performance validation.
The optical throughput of a monochromator depends on the source, the slit height,
•
the collected solid angle, the transmission factor of the optics, and the convolution of
the entrance and exit slit widths (dispersion). The light gathering capacity (LGC) is
defined by:
LGC
height mm
dispersion
mm
mm
slit
=
(
)
( / #) *
f
2
(5.11)
5.4.4 Polarization Effects
Polarization effects in monochromators and other optical components can present significant difficulties in the overall operation and calibration of fluorimeter systems. At the same
time, the introduction of polarizing elements such as polarizing filters or Glan–Thompson
or Glan–Taylor polarizing optics are essential to the measurement of fluorescence polarization or anisotropy.
Reflection diffraction gratings are well described (www.horiba.com, www.newport.
com) and can exhibit both strong and complex polarization effects. At certain wavelengths
the grating may exhibit diffraction efficiencies in the S- or P-polarization planes. On average, this has little effect on the overall power transmitted by the monochromator for generally unpolarized light inputs but the monochromatic beam that exits the monochromator
will, to some extent, be partly polarized, the extent of which can be strongly wavelength
dependent. These effects lead to spectral shifts, signal loss, and a whole range of other
misleading artefacts in the spectral signal. Figure 5.14 shows two sets of typical diffraction
grating efficiency curves, from two master gratings, for 1200 g mm
–1
and 500 nm blazed
gratings with polarization angles for 45 degrees (upper boxes) and also S- (perpendicular)
and P- (parallel) planes (lower boxes).
Nondispersive elements such as filters are also widely used in all of the optical spectroscopy methods, especially fluorescence measurements, and are based on either absorption
or interference. Filters are commercially available for wavelengths above 200 nm and come
in many forms, some of which are bandpass, cutoff, heat-absorbing, heat-reflecting, etc.
The most common types are:
Bandpass filters of the interference type are defined by the bandpass wavelength and the
•
width of the bandpass. It is common to use such filters to select a wavelength; that is,
166
However, this type of approach is not realistic to the normal use of monochromators
as broad-band tunable light sources or broad-band detection system. Typically, stray light
is very difficult to measure, as it strongly depends on the wavelength, bandpass used, and
the type of source. A further discussion of this important point is made in Section 5.4.8 on
instrument performance validation.
The optical throughput of a monochromator depends on the source, the slit height,
•
the collected solid angle, the transmission factor of the optics, and the convolution of
the entrance and exit slit widths (dispersion). The light gathering capacity (LGC) is
defined by:
LGC
height mm
dispersion
mm
mm
slit
=
(
)
( / #) *
f
2
(5.11)
5.4.4 Polarization Effects
Polarization effects in monochromators and other optical components can present significant difficulties in the overall operation and calibration of fluorimeter systems. At the same
time, the introduction of polarizing elements such as polarizing filters or Glan–Thompson
or Glan–Taylor polarizing optics are essential to the measurement of fluorescence polarization or anisotropy.
Reflection diffraction gratings are well described (www.horiba.com, www.newport.
com) and can exhibit both strong and complex polarization effects. At certain wavelengths
the grating may exhibit diffraction efficiencies in the S- or P-polarization planes. On average, this has little effect on the overall power transmitted by the monochromator for generally unpolarized light inputs but the monochromatic beam that exits the monochromator
will, to some extent, be partly polarized, the extent of which can be strongly wavelength
dependent. These effects lead to spectral shifts, signal loss, and a whole range of other
misleading artefacts in the spectral signal. Figure 5.14 shows two sets of typical diffraction
grating efficiency curves, from two master gratings, for 1200 g mm
–1
and 500 nm blazed
gratings with polarization angles for 45 degrees (upper boxes) and also S- (perpendicular)
and P- (parallel) planes (lower boxes).
Nondispersive elements such as filters are also widely used in all of the optical spectroscopy methods, especially fluorescence measurements, and are based on either absorption
or interference. Filters are commercially available for wavelengths above 200 nm and come
in many forms, some of which are bandpass, cutoff, heat-absorbing, heat-reflecting, etc.
The most common types are:
Bandpass filters of the interference type are defined by the bandpass wavelength and the
•
width of the bandpass. It is common to use such filters to select a wavelength; that is,