[RASMB] question on wavelength calibration of AUC

[Jan Komárek]

Dear RASMB members, 
I have a question related to wavelength calibration/accuracy of ABS optics.
There is suspicion that the calibration on our XL-I may be few nm out. 

When looking at the peaks in wavelenght scan (emission spectrum of the lamp),
position of the 229 and 529 nm peaks are fine and within the specifications.
But when I use certified OD check solution (potassium nitrate), I can see the
maximum at 297 nm (instead of declared 302 nm). Also, when I perform the
wavelength scan of the protein sample, the maximum is not 280 nm, but 274-275
nm (and therefore I get lower signal for protein samples than I would
expect). 

We did some other tests after discussions with Beckman technicians. The
reproducibility of the monochromator seems to be good (deviations within 1 nm
between several wavelength scans performed in a row) and the stray light check
was also OK. The details are enclosed. Unfortunatelly we don´t have holmium
oxide cell to independently verify the wavelengths.
I am aware of the limitations in wavelength accuracy on XL-A/I and know that
the wavelenghts will never be perfect, but I would expect that both
wavelenght/intensity check and OD check should give the same results...

Has anybody seen something similar or is anybody able to explain the
discrepancy? Any suggestion/comment would be appreciated. Thank you very much
and have a nice day :)

With kind regards
Jan Komarek




-- 
Jan Komarek
Central European Institute of Technology (CEITEC)
Masaryk University
Kamenice 5, A4/3.13
625 00 Brno, Czech Republic
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