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Absorption/Emission Spectra Of Rhodamine B In Ethanol

A project log for DAV5 V3.01 Raman Spectrometer

The only thing worth doing, is the thing worth doing right!

david-h-haffner-srDavid H Haffner Sr 01/05/2017 at 11:590 Comments

Log#41; Further qualifying the UV/VIS functionality of the DAV5 V3 Raman spectrometer by imaging a sample of Rhodamine B (Rhodamine dyes are used extensively in biotechnology applications such as fluorescence microscopy, flow cytometry, fluorescence correlation spectroscopy and ELISA.)

In fig.1, To the left is my reference spectra (Rhodamine B in Ethanol from the Oregon Medical Laser Center/Using a SpexFluoroMaxII Fluorometer) To the right, is my sample of Rhodamine B in Ethanol using my DAV5 V3 Raman spectrometer.

The reason the FWHM values are different between the two spectra are, because the SpexFluoroMax II, uses a Xenon lamp as the broadband light source and a CCD detector. The DAV5 V3 uses a JDEPC-05 cmos 0.25'' board level detector and a 532nm DPSS laser source.

Note that this does NOT in any way diminish the data, as you can see that the spectral shapes are nearly identical.

In Fig.2 above, this is an absorption and emission spectra taken with the DAV5 V3 spectrometer. The absorption spectra was obtained using a Solux lamp/50W/12vdc/5A/4700K. The emission spectra was obtained using an Aries 532nm DPSS green laser/150mW/3vdc.

Samples were prepared in 1cm pathlength quartz cells with absorbance less than 0.1 at the excitation and all emission wavelengths to uniformly illuminate across the sample, and to avoid the inner-filter effect. The dark counts were subtracted and the spectra were corrected for wavelength-dependent instrument sensitivity.

References:

http://omlc.org/spectra/PhotochemCAD/html/009.html - rhodamine b in ethanol

http://www.turnerdesigns.com/t2/doc/appnotes/998-5111.pdf - tracer dye prep

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