I want to demonstrate that FITC is present in a powder obtained by the micronization method.
Firstly, FITC is orangish in color. If you shine a UV light on it, it will glow greenish. If you dissolve some in a neutral or alkaline pH buffer and take an absorbance or fluorescence spectrum, the spectrum will be that of fluorescein. It will react with primary amines, which you will be able to judge by thin layer chromatography or other chromatography method.
It's not clear to me what really you wish to know? How to measure the intensity of fluorescence or to identify a substance? To measure fluorescence you just need a spectrofluorometer. Identification of a substance is a tricky problem. A simple way is to use HPLC or TLC. Spectroscopic techniques are usually less selective. Regarding FITC, you can use both fluorescence and absorption. The absorption spectrum has a maximum at around 530 nm, depending on pH of a solution. Fluorescence of FITC includes two types of emission: conventional fluorescence and phosphorescence. So, if I were you I would use the latter emission, because not many dyes are capable of emitting phosphorescence. However, there is a complication. Phosphorescence is strongly quenched by oxygen at room temperature. Thus, it is necessary to remove oxygen from your sample or slow down the diffusion rate of oxygen. You can do it if you freeze your sample. As a solvent, it would be better to use glass-forming or viscous liquids, like 70/30 (v/v) glycerol/water mixture to keep your sample transparent during freezing. Another aspect is excitation of your sample. All dyes absorb in the UV region, therefore excitation of a sample with a UV source is not selective. It would be more correct to use a light source with a wavelength around 530 nm where FITC has its selective absorption band. When I worked with EITC (another xanthene dye), I used a Nd-YAG laser with excitation at 532 nm. So, if you do this, you will be able to see the long-lived luminescence (with lifetime in the range of microseconds-seconds) at about 600 nm (greenish light). Therefore, using the selective excitation and emission wavelengths as well as the long-living luminescence will help you prove that the tested dye is FITC. Of course, it is impossible to describe all aspects you have to take into account preparing your particular experiment.
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