NRU :: Browsing by Author "Magin, Richard L."

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A Fiber-optic System for Measuring Single Excitation-Dual Emission Fluorescence Ratios in Real Time
(Biotechnology progress, 1992) McCarthy, John F.; Magin, Richard L.; Kisaalita, William S.; Slininger, Patricia J.
The development and subsequent evaluation of a fiber‐optic system for measuring single excitation—dual emission fluorescence ratios in real time is described. The design of the flashlamp excitation source, optics, electronics, and computer software is discussed. The dual emission pH sensitive fluorophore 1, 4‐dihydroxyphthalonitrile (1, 4‐DHPN) was used to test the performance of this system. The flexible design of this modular system permits the use of other single excitation—dual emission fluorophores by simply changing the appropriate optical filters. Upon a single 340–380‐nm excitation, pH‐sensitive emissions were monitored at 488 nm and 434 nm. The ratio of these emissions (488/434) was then computed in real time, for a 2 mM solution of 1, 4‐DHPN, while the pH was titrated over the range 5–9. The nonlinear, system‐dependent, calibration curve of pH versus the ratio of emission wavelengths was empirical fit by a fourth‐order polynomial (r2 = 0.995). Reliable pH measurements in the range 6–8 were obtained using concentrations of 1, 4‐DHPN as low as 50 μM. The standard deviation of pH measurements using a 1 mM solution of 1, 4‐DHPN, near neutral pH, was found to be approximately 0.1 pH unit