spectrofluorometer

A spectrofluorometer, or fluorescence spectrophotometer, is an analytical instrument used to measure the fluorescence emitted by a sample after it is excited with light. It provides qualitative and quantitative information about fluorophores, their environment, and their concentration by recording emission spectra and, in some cases, excitation spectra.

A typical instrument consists of a light source, an excitation monochromator to select the excitation wavelength, an emission monochromator to resolve the emitted light, a sample compartment, and a detector such as a photomultiplier tube or a photodiode array. Additional components may include optical filters and polarization optics. The system is designed to minimize stray light and maximize signal-to-noise.

Measurement modes include emission scans (varying emission wavelength at a fixed excitation), excitation scans (varying excitation wavelength while monitoring emission at a fixed wavelength), and, in advanced models, time-resolved fluorescence to measure lifetimes. Data are usually plotted as fluorescence intensity versus wavelength and can be corrected for instrument response or background.

Applications span chemistry and biochemistry, including fluorophore characterization, protein and nucleic acid labeling assays, receptor–ligand binding studies, environmental analyses using fluorescent tracers, and pharmaceutical quality control. Fluorescence quantum yield and sensitivity are key performance metrics.

Advantages include high sensitivity, wide dynamic range, and relatively straightforward sample handling. Limitations include photobleaching, inner-filter effects at high absorbance, and the need for careful calibration with standards. Proper solvent choice, temperature control, and routine instrument maintenance improve accuracy and reproducibility.