Influence of tissue fluorescence measurement and imaging by autofluorescence of substrata
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Date
2015-08
Journal Title
Journal ISSN
Volume Title
Publisher
Journal of Applied Spectroscopy
Abstract
Polymers, glass and fi ber reinforced materials are increasingly being used in photophysical devices. The inherent
fl uorescence of devices made of these materials may itself become important during fl uorescence detection. It may
interfere with or even negate the low fl uorescence signals emitted from fl uorophores molecules of interest especially
when the fl uorophores exist in microvolume samples. Such effects are signifi cant when tissue micro-sections placed
on substrata of high background fl uorescence.
Using different excitation wavelengths, fl uorescence spectral and imaging studies were carried out to examine
the effect of background fl uorescence of microslides made of various substrata such as glass, polycarbonate and
aluminium on autofl uorescence of tissue sections. With decreasing wavelength of excitation, all substrata showed
increased autofl uorescence. Glass showed least inherent fl uorescence and aluminium showed more refl ectance
than autofl uorescence. After continuous exposure to blue and UV light, all substrata showed irreversible decreased
autofl uorescence. During spectrofl uorimetric studies, a defi nite blue shift was observed in the autofl uorescence of tissue
sections placed on microslides made of each of the materials. This suggests an interference of tissue autofl uorescence
by background fl uorescence emitted by the substratum. This may lead to false positive or negative reporting of the
fl uorophores, particularly important while analyzing micro-sections of biological samples. Microscopic imaging
did not show any background fl uorescence for glass substratum with blue light. However, with UV light, glass
also showed background fl uorescence during imaging. Other substrata showed strong background fl uorescence
or refl ectance with both blue and UV light. This study has the potential for accurate quantifi cation of fl uorescence
spectral and decay and improved fl uorescence imaging using the photobleaching effect.
Description
Keywords
auto-fl uorescence, substratum, photobleaching, blue shift.
Citation
Santhosh Kumar B, Sandhyamani S, Nazeer SS, Jayasree RS. Influence of tissue fluorescence measurement and imaging by autofluorescence of substrata. Journal of Applied Spectroscopy. 2015;82(3):494-501