Design considerations for a fluorophotometer for ocular research

Abstract

Fluorometric studies have been made with modified slit-lamp microscopes. These instruments offered considerable versatility in modifying experimental parameters, but were seldom optimized in terms of detector and illuminator apertures, filter band-pass characteristics, detection system, light-source stabilization, or optical resolution. Improper control of these parameters can result in artifacts which override useful information.

A new fluorophotometer for ocular research has been designed that utilizes an optical system, which eliminates the need for a contact lens. An effort was made to optimize both the axial resolution and the minimum detectable signal.

The characteristics of standard barrier filters and excitation filters were examined and compared with photomultiplier sensitivity curves. It was necessary to use double filters in order to reduce artifacts due to excitation light that was not rejected by the barrier filter.

In order to optimize the signal-to-noise ratio, a photon counting detection system was used. A dark current equivalent to 3 × 10⁻¹⁰ g/ml allows a useful detection range of 1 × 10⁻¹⁰ g/ml.