| 摘要 |
A quantitative fluorescence technique is presented as "differential real-time fluorescence spectroscopy" (DRFS), pursuant to which one can simultaneously make time-resolved measurement of fluorescence changes in two cuvette samples (e.g., experimental and control), wherein the fluorescence is induced by excitation light, and wherein both samples have been identically prepared with a selected ratiometric dye. The ratiometric dye may either be of excitation-shifted variety or of emission-shifted variety. With DRFS, it is possible to monitor the response of specimen cells to a chemical, biological or physical agent, in the absence and in the presence of a selected experimental variable, and to determine both of these responses in real time, i.e., at the same time as the measurements are being made. The invention is therefore ideally suited to monitor response from experimental stimuli that are expected to induce only relatively small cellular changes during the typical time course of a fluorescence measurement, namely, 20 minutes or less. The reason for this capability is that DRFS eliminates problems attributable to inherent biological variability associated with preparations of living cells, such as known and unknown (e.g., time-dependent) changes in base-line biological activity during the storage or handling of prepared cells, prior to and awaiting a particular experiment.
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