UV Fluorescent imaging systems

Fluorescent imaging is widely used in many branches of the life sciences. Biological samples are selectively stained using dyes that bind to parts or molecules of interes and sing fluorescent stains can give higher sensitivity compared to coloured stains. Typically, imaging systems comprise:

• Excitation source lamp (mercury discharge, xenon, laser etc.). This is chosen to maximise power in the excitation band of the fluorophore and, ideally, minimum power in its emission band.
• Optical filter to select the excitation wavelengths suitable for the fluorophore(s) being imaged.
• Excitation delivery optics. It is important to deliver the excitation to the sample at high power and with good uniformity over the field of view
• Sample holder.
• Optical emission filter. For high sensitivity it is essential that very little of the exication wavelengths reach the detector. By design of the filters, the passband of one filter can be arranged to coincide with a high-attenuation stop band of the other filter.
• Imaging optics and sensor. Typically this is a high resolution CCD camera, cooled to minimise dark current and noise.

Three development projects have been undertaken to date. The first was a stand-alone imaging system which offered a viewing area of up to 250mm by 250mm, but could zoom onto much smaller samples. Illumination was through the sample (trans-illumination), with options to swap quickly between different visible and UV excitation wavelengths. Two further projects integrated the imaging system with sampling robots so that sample manipulation can be directed by on-line images. One of the imaging systems used low pressure mercury tubes for visible or UV trans-illumination whereas the other had a xenon arc lamp and a system of interference filters for top (epi-) illumination. In the latter system, filter swapping was under computer control, integrated with the robot, so that multiple fluorophores could be imaged in the same sampling process.

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