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ation with macromolecules, and excit ed-state complex formation can be studied for subsets of the fluorescent components that fall within a given lifetime range. Suppression of interfering sig nals. The minimization of unwanted signals is inherent to the concept of fil tering. In time-resolved experiments, gated detection can be used to reduce the contribution of short-lived inter fering signals to the measured intensi ty. In phase-resolved experiments, in terfering signals that are either longeror shorter-lived than the desired signal can be relatively suppressed in several ways: by setting the detector phase to be out-of-phase with the interfering
signal, by using a modulation frequen cy at which the interfering signal is greatly demodulated, or by combining detector phase and modulation fre quency. Longpass filters in the time domain are ideal for certain applications in which the desired signal is much long er-lived than the background signal. For example, scattered light interfer ences can be greatly reduced, improv ing signal-to-scattering ratios by more than twofold for short-lived (τ = 0.7 ns) fluorescence and by sixfold for longerlived fluorescence signals (10). Detec tion limits can also be improved for compounds with very long-lived emis sion, such as lanthanide chelates (r =
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LABCONCO 145 MILTON ROY/APD 37 NEYTECH/J.M. NEY . . . . 101-175 OMEGA IFC PHARMACIA LKB 65 RUDOLPH RESEARCH 113 SHIMADZU 53 SWAGELOK 76-77 UICINC 101 VALCO 141 WHEATON 99,119
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Figure 5. Fluorescence EEMs of pooled serum. (a) Steady-state EEM of a 20-fold dilution, showing scattered light peak (extending well off-scale) at the rear and second-order scattered light in the foreground; (b) PREEM of the same solution, collected at 30 MHz, with scattered light signal suppressed; and (c) PREEM of a fivefold dilution of the serum at 30 MHz, corresponding to the right rear quadrant of the EEMs in (a) and (b). (Adapted with permission from Reference 14.)
844 A · ANALYTICAL CHEMISTRY, VOL. 6 1 , NO. 14, JULY 15, 1989
