Evaluation of Low Noise Objective Lenses for Detection of Weak Fluorescence

Extract

The optimal method for evaluating fluorescence microscope has not been proposed. In this study we developed two quantitative methods using biological and non-biological materials as standard specimens. Based on these methods, a new water immersion lens was tested and its high sensitivity to fluorescence was demonstrated.

First, particles of a fluorescent glass (Luminass-1; Sumita Optical Glass) were used for the standard specimen. Particles with a few micrometer diameter were embedded in silicon oil on a quartz slide glass covered with a quartz glass slip and then observed. Under the fluorescence microscope, the spectra of emission light from the excited area with or without the particle were measured by a monochromator and a photomultiplier tube (PMT). The signal-to-background ratio (S/B) was calculated from these spectra (Fig. 1, Table 1). This specimen showed a significant advantage due to less photobleaching that permits a stable and long observation time.