Biomedical Engineering Reference
In-Depth Information
Fig. 9.8 Configuration of a
typical fluorescence imaging
system
9.3.2.1
Light Sources
The most important requirement of the light source in fluorescence imaging is a
close match of the spectrum to the excitation wavelength of the fluorochrome to
achieve high brightness and high contrast images. The wavelength of the light
source determines which type of fluorophore can be excited and how well that
fluorophore can be excited. The wavelength away from the peak of the excitation
spectrum may produce background noise that can overshadow the fluorescence
signal of interest. The wavelength is also the major factor in determining the probed
depth, given that the penetration depth of light in tissue strongly depends on the
wavelength.
Light source intensity is the other major factor in fluorescence imaging. Most
fluorophores have low quantum yields; therefore, powerful light sources are needed
to excite enough fluorescent light to be detected by the eye or other detectors.
There are two types of light sources used in fluorescence imaging: noncoherent
and coherent sources. Noncoherent light sources are usually broadband; these
are typically used in spectroscopy and area imaging systems. The most common
noncoherent light sources are mercury, xenon, and metal halide arc lamps.
The light intensity of a mercury (HBO) lamp is not uniform from UV to near-IR
light, with peaks of intensity at 313, 334, 365, 406, 435, 546, and 578 nm; in between
the peaks, the energy of the light source is very low. HBO lamps are suitable for the
applications that need blue or UV light to excite fluorescence.
Xenon arc (XBO) lamps have a relatively flat emission spectrum without
emission peaks across the visible spectrum. The uniform emission levels and lower
fluctuations make XBO lamps better suited to applications in ratio imaging and
other quantitative applications. XBO lamps also have the advantage that they exhibit
stable emission intensity over time.
Mercury-xenon arc lamps have the best characteristics of both xenon and
mercury lamps. The spectral distribution of a mercury-xenon lamp includes a
continuous spectrum from the ultraviolet to infrared and a strong mercury line
spectrum. It has a bluish-white spectrum and extremely high UV output.
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