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Phosphorescence is the emission of light, in which the excited state electron has the same spin orientation as the ground state electron. This transition is a forbidden one and hence the emission rates are slow (10 3 - 10 0 s -1 ). So the phosphorescence lifetimes are longer, typically seconds to several minutes, while the excited phosphors slowly returned to the ground state. Phosphorescence is still seen, even after the exciting light source is removed. Group 12-16 semiconductor quantum dots exhibit fluorescence properties when excited with ultraviolet light.


The working schematic for the fluorometer is shown in [link] .

Schematic of fluorometer.

The light source

The excitation energy is provided by a light source that can emit wavelengths of light over the ultraviolet and the visible range. Different light sources can be used as excitation sources such as lasers, xenon arcs and mercury-vapor lamps. The choice of the light source depends on the sample. A laser source emits light of a high irradiance at a very narrow wavelength interval. This makes the need for the filter unnecessary, but the wavelength of the laser cannot be altered significantly. The mercury vapor lamp is a discrete line source. The xenon arc has a continuous emission spectrum between the ranges of 300 - 800 nm.

The diffraction grating and primary filter

The diffraction grating splits the incoming light source into its component wavelengths ( [link] ). The monochromator can then be adjusted to choose with wavelengths to pass through. Following the primary filter, specific wavelengths of light are irradiated onto the sample

Sample cell and sample preparation

A proportion of the light from the primary filter is absorbed by the sample. After the sample gets excited, the fluorescent substance returns to the ground state, by emitting a longer wavelength of light in all directions ( [link] ). Some of this light passes through a secondary filter. For liquid samples, a square cross section tube sealed at one end and all four sides clear, is used as a sample cell. The choice of cuvette depends on three factors:

  1. Type of solvent - For aqueous samples, specially designed rectangular quartz, glass or plastic cuvettes are used. For organic samples glass and quartz cuvettes are used.
  2. Excitation wavelength – Depending on the size and thus, bandgap of the Group 12-16 semiconductor nanoparticles, different excitation wavelengths of light are used. Depending on the excitation wavelength, different materials are used ( [link] ).
    Cuvette materials and their wavelengths.
    Cuvette Wavelength (nm)
    Visible only glass 380 - 780
    Visible only plastic 380 - 780
    UV plastic 220 - 780
    Quartz 200 - 900
  3. Cost – Plastic cuvettes are the least expensive and can be discarded after use. Though quartz cuvettes have the maximum utility, they are the most expensive, and need to reused. Generally, disposable plastic cuvettes are used when speed is more important than high accuracy.
A typical cuvette for fluorescence spectroscopy.

The cuvettes have a 1 cm path length for the light ( [link] ). The best cuvettes need to be very clear and have no impurities that might affect the spectroscopic reading. Defects on the cuvette, such as scratches, can scatter light and hence should be avoided. Since the specifications of a cuvette are the same for both, the UV-visible spectrophotometer and fluorimeter, the same cuvette that is used to measure absorbance can be used to measure the fluorescence. For Group 12-16 semiconductor nanoparticles preparted in organic solvents, the clear four sided quartz cuvette is used. The sample solution should be dilute (absorbance<1 au), to avoid very high signal from the sample to burn out the detector. The solvent used to disperse the nanoparticles should not absorb at the excitation wavelength.

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