Pre-Assignment

1. Mention three different ways by which the fluorophore concentration can affect the fluorescence spectral characteristics.
2. Mention two ways by which one can avoid inner filter effects.
3. Explain how fluorophores with large Stokes shifts are less sensitive to the inner filter effects.
4. Mention the geometrical as well as the mathematical reason for which the fluorescence intensity of a fluorophore is proportional to the concentration over only a limited range of absorbances. (Hint: Mathematical: IF = kφF I0 (1 e-2.3εcl ) and (1- e2.3A) = 2.3 A, when A << 1)
5. What kinds of illumination geometries are useful for fluorescence studies of optically dense samples or highly scattering solutions?
6. What kind of illumination geometry should one use to minimize the excitation inner filter effect?

Post-Assignment

1. Why should one start the measurement with the lowest concentration first and go on to the next higher concentration while doing the measurement?
2. Why should one always rinse the cuvette with the experimental solution prior to filling up of the cuvette with the experimental solution?
3. What kind of illumination geometry was used in this experiment?
4. Why does the shorter wavelength part of anthracene fluorescence get attenuated unlike its long wavelength part?
5. Why does the fluorescence attenuation increase with the increasing anthracene concentration?