Page 56 - Fundamentals of Light Microscopy and Electronic Imaging

FILTERS FOR ADJUSTING THE INTENSITY AND WAVELENGTH OF ILLUMINATION 39

them. These filters must be inserted into the beam with the reflective surface facing the
light source. They can, however, be cheaper and thinner, and are the filter of choice for
use with lasers.

Colored Glass Filters

Colored glass filters are used for applications not requiring precise definition of trans-
mitted wavelengths. They are commonly used to isolate a broad band of colors or as
long-pass filters to block short wavelengths and transmit long ones. Colored glass filters
contain rare earth transition elements, or colloidal colorants such as selenide, or other
substances to give reasonably sharp transmission-absorption transitions at a wide range
of wavelength values across the visual spectrum. Since colored glass filters work by
absorbing quanta of nontransmitted wavelengths, they can be heat sensitive and subject
to altered transmission properties or even breakage after prolonged use. However, as the
absorbent atoms are contained throughout the glass and are not deposited on its surface,
colored glass filters offer major advantages: They are less resistant to physical abrasion
and chemical attack from agents contained in fingerprints and other sources, and their
optical performance is not sensitive to the angle of incidence of incoming rays of light.
Colored glass filters are also less expensive than interference filters and are generally
more stable and long-lived.

Interference Filters

Interference filters often have steeper cut-in and cut-off transmission boundaries than col-
ored glass filters and therefore are frequently encountered in fluorescence microscopy
where sharply defined bandwidths are required. Interference filters are optically planar
sheets of glass coated with dielectric substances in multiple layers, each /2 or /4 thick,
which act by selectively reinforcing and blocking the transmission of specific wavelengths
through constructive and destructive interference (Fig. 3-6). Bandpass filters transmit a
limited range of wavelengths that experience reinforcement through constructive interfer-
ence between transmitted and multiple reflected rays; wavelengths that do not reinforce
each other destructively interfere and are eventually back-reflected out of the filter.
Interference filters contain layers of dielectric substances, electrically nonconduc-
tive materials of specific refractive index, typically optically transparent metal salts such
as zinc sulfide, sodium aluminum fluoride (cryolite), magnesium fluoride, and other sub-
stances. In some designs semitransparent layers of metals are included as well. The inter-
face between two dielectric materials of different refractive index partially reflects
incident light backward and forward through the filter, and is essential for constructive
interference and reinforcement. The wavelength that is reinforced and transmitted
depends on the thickness and refractive index (the optical path) of the dielectric layers.
The coatings are built up in units called cavities, with 1 cavity containing 4 or 5 alternat-
ing layers of dielectric salts separated by a spacer layer (Fig. 3-7). The steepness of the
transmission boundary and the definition of filter performance are increased by increas-
ing the number of cavities. An 18-cavity filter may contain up to 90 separate dielectric
layers. The deposition of salts is performed by evaporation of materials in a computer-
controlled high-vacuum evaporator equipped with detectors for optical interference,
which are used to monitor the thicknesses of the various layers. The final layer is covered