Table 1.

Characteristics of the Internal Reference

	Red Slide* + NDF† (1.3 DU)
Dimensions, mm	7 ×5 ×1.3
Peak emission, nm	590
Half-height points, nm	570–615
Change in excitation spectrum from 480 to 490 nm, %	+12
Fluorescence decay to 5% of peak, ns	29‡
Change in fluorescence after a 300-h exposure with 1.3 mW/cm2 of 470–500-nm blue light, %	+0.8 ± 0.05§
RCF
HRA2	260 (515)¶
Spectralis	231
Sampling Area	180 × 23 pixels

^*Developed and tested for stability by Ping Chin Cheng (SUNY, Buffalo, NY). The fluorophore is Texas Red dye with other proprietary compounds embedded in a plastic matrix (Microscopy/Microscopy Education, McKinney, TX).

^†Neutral-density filter (Wratten; Kodak, Rochester, NY) optically cemented to the slide. This method provides an attenuation of 10^2×1.3 = 398, since both the excitation and emission are affected.

^‡The decay time must be short enough to ensure that sufficient fluorescence is detected before the horizontal scanner moves to neighboring areas (pixel clock: 100 ns in high-speed mode).

^§The test irradiance was 14× higher than the irradiance in the intermediate plane and 280× higher (14 × 10^1.3) than the irradiance on the fluorescent material.

‖NAn early version of the internal reference, used here in a few tests, used a lower NDF of 1.1 DU; the RCF was 515.