Previously when I run my samples I got good results with 3D fluorescence spectrometer( LS-55 ) but now I cant get the same results again and there are some scattering lines and the place of excitation and emission wavelength are changed . I don't know if there is something wrong with instrument or my method setting. The attached file is one of my sample's (EEM) before and after the changes.
first you can check , sample reply shows some emission,
your first sample also has some scattering but molecular fluorescence was high enough
but second spectra , there is weak emission from your molecule that's why your emission can not seen in matrix
Thanks Dear Karuppannan Senthilkumar, but my question is that why suddenly it happens?Is it something wrong with instrument or software?Why in the second one we have weak emission from molecules?
Hi there,
the second EEM shows only second order straylight and some Raman scattering, which are both artifacts from the spectrometer optical system. These artifacts show thath at least the system is able to measure "something". EEMs look like this, when no emitting species was present! There is a multitude of possible causes. Either:
- no cuvette was inserted
- an empty cuvette was inserted
- the cuvette contained only solvent
- the emitting species is no longer there (due to decomposition caused by photobleaching, or chemical reaction, or pH change, or whatever)
To rule out any instrument/software related problem, you might try a "known" (stable, well fluorescing )fluorescent solution, like fluorescein in NaOH or Rhodamin 6G in ethanol...
yes you are right , photobleaching may be one of the reason for this,
so check the absorption spectra of that molecule after the measurement
then you could know where is the problem
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