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Updated: Jun 6, 2025

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Optimising Shifted Excitation Raman Difference Spectroscopy (SERDS) for application in highly fluorescent biological

H Sheridan1, A P Dudgeon1,2,3, J C C Day3

  • 1Biomedical Physics, Department of Physics and Astronomy, University of Exeter, Exeter, EX4 4QL, UK.

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Summary

Shifted excitation Raman difference spectroscopy (SERDS) overcomes fluorescence and silica signals in fiber optic Raman spectroscopy. An optimal 2.4 nm wavelength shift was identified for analyzing biological samples, including human lymph nodes.

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Area of Science:

  • Biophotonics
  • Spectroscopy
  • Medical Diagnostics

Background:

  • Fiber optic Raman spectroscopy enables in vivo molecular analysis but is limited by tissue fluorescence and silica probe signals.
  • High fluorescence can cause CCD etaloning, complicating data analysis.
  • Shifted excitation Raman difference spectroscopy (SERDS) is a technique to mitigate these interferences.

Purpose of the Study:

  • To determine the optimal wavelength shift for SERDS to reconstruct narrow and broad peaks in biological samples.
  • To evaluate the performance of SERDS with varying excitation wavelength shifts.
  • To demonstrate the feasibility of SERDS using a fiber optic probe for ex vivo lymph node analysis.

Main Methods:

  • Utilized an 830 nm excitation wavelength with seven different shifts (0.4–3.9 nm).
  • Investigated peak widths relevant to biological Raman spectra (0.41–3.25 nm or 6–47 cm-1).
  • Applied the optimized SERDS method to ex vivo human lymph node samples using a fiber optic probe.

Main Results:

  • An optimal wavelength shift of 2.4 nm was identified for SERDS.
  • This shift effectively removed fluorescence and etaloning artifacts while preserving biological Raman signals.
  • Successful ex vivo measurement of human lymph nodes demonstrated the technique's practical application.

Conclusions:

  • SERDS with an optimized wavelength shift significantly enhances the utility of fiber optic Raman spectroscopy for biological tissue analysis.
  • The 2.4 nm shift provides optimal spectral reconstruction for diverse biological Raman peaks.
  • This approach holds promise for improved disease diagnostics through non-invasive molecular analysis.