Selection Of Barrier Filter (Ba Filter) - Nikon Eclipse 80i Instructions Manual

Brightness of fluorescent image
Generation of self-fluorescence
Degree of color fading

Selection of barrier filter (BA filter)

18.2
The barrier filter allows only fluorescent light emitted by the specimen to pass, blocking
excitation light. This allows viewing of a fluorescent image without excess illumination (dark
background).
There are two types of barrier filters: LP filters block all light below a certain wavelength but
pass all light of longer wavelengths. BP filters pass only light of a certain waveband, blocking all
other light. Use the filter type appropriate for your intended purpose.
LP filter (long-pass filter)
LP filters block all light below a certain wavelength
but pass all light of longer wavelengths. This border
wavelength is called a cut-on wavelength.
(1) For specimens dyed with a fluorescent
pigment in which the fluorescent waveband
and excitation waveband (light that the
specimen absorbs in order to emit fluorescent
light) are very close, select a barrier filter
with the shortest cut-on wavelength
permitted by performance requirements for
efficient fluorescent microscopy. If the cut-on
wavelength is long, excitation light and
fluorescent light will be entirely distinct,
tending to darken the background of
fluorescent images. However, recent
advances in filter performance have resulted
in increased use of filters of short cut-on
wavelengths.
(2) For multiple-dye specimens, use an LP filter
for microscopy of fluorescent images of all
dyes. Note that a combination involving an
ordinary dichroic mirror, an excitation filter,
and an LP-filter-type barrier filter will be
incapable of exciting dyes that emit
long-wavelength fluorescent light – for
example, TRITC in the case of FITC and
TRITC. This will result in very dark TRITC
fluorescent images. For such cases, we
recommend using multiband filters.
Individual Operations
18 Selecting Fluorescent Filters
Narrow
Dark
Low
Low
32
EX filter bandwidth
Waveband
emitted by
FITC
Wavelength
Both the FITC and TRITC images are visible.
Wide
Light
High
High
LP520
Waveband
emitted by
TRITC