I made serious concentrations of bovine serum albumin 200, 150,100, 50, 30, 25, 20, 15, and 10 uM. Once I measured the fluorescence intensity, I found that there was a gradual increase in the fluorescence intensity. On the other hand, I conjugate that Protein with AuNPs and the fluorescence intensity recorded at different values, whereas, AuNPs alone was recorded at very low value as a comparison with the conjugated ones. Any Ideas about this differences and if you know any methods to count that protein molecules present onto the surface of gold nanoparticles?
Many thanks in advance
The first answer is straightforward. In steady-state emission, you excite the molecules (BSA for you) and sums up the radiative emission for a period of time. The more molecules you have, the more molecules will be excited, and the more fluorescence you will get. Upon conjugating the protein with AuNPs, you open nonradiative pathways, such as FRET, that dramatically influence the sluorescence you measured.
It is quite natural that emission intensity increases with the increasing concentration of fluorophores. When you are increasing the BSA concentration, the amount of Trp increase and that affect the emission increment.
The second part is not clear. You need to mention the excitation and emission wavelength.
Fluorescence intensity can increase with increasing fluorophore concentration. Typically, in dilute solutions the fluorescence intensity is directly proportional to the fluorophore concentration. However, in concentrated solutions, this situation may not be entirely linear. Also at higher concentrations certain fluorophores will tends to aggregate and quench the fluorescence. This phenomena is called the aggregation-caused quenching (ACQ).
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