Time‐gated interferometric detection increases Raman scattering to fluorescence signal ratio in biological samples

Abstract

Attainable levels of signal-to-background ratio (SBR) in Raman spectroscopy of biological samples is limited by the presence of endogenous fluorophores. It is customary to remove the ubiquitous fluorescence background using postacquisition data processing. However, new approaches are needed to reduce background contributions and maximize the fraction of the sensor dynamical range occupied by Raman photons. Time-resolved detection using pulsed lasers and time-gated

Multispectral label‐free Raman spectroscopy can detect ovarian and endometrial cancer with high accuracy

“…Background removal typically relies on the relatively smooth fluorescence background spectrum compared to sharper Raman features, by utilizing hyperspectral detection combined with polynomial curve fitting. However, other techniques are currently being developed that could allow single bands to be imaged, for example, early-photon time-gated detection to isolate nonresonant from resonant phenomena, that is, Raman scattering from fluorescence [18].…”

Open-sourced Raman spectroscopy data processing package implementing a baseline removal algorithm validated from multiple datasets acquired in human tissue and biofluids

“…This is done by limiting light detection to non-resonant interaction phenomena with relaxation time scales occurring on sub-nanosecond timescales, thereby excluding contributions from resonant phenomena such as fluorescence. 36 …”

“…This is done by limiting light detection to non-resonant interaction phenomena with relaxation time scales occurring on sub-nanosecond timescales, thereby excluding contributions from resonant phenomena such as fluorescence. 36 The common denominator of most of these Raman spectroscopy methods is that they lend spectra composed of up to 1000 to 2000 intensity bins (e.g., vectors) that require post-acquisition processing data treatment to isolate the inelastic scattering contribution associated with the interrogated material. This is essential because the inelastic scattering (i.e., Raman scattering) contribution is typically orders of magnitude smaller when compared to endogenous fluorescence from tissue biomolecules (e.g., collagen, elastin, NADH, and FAD).…”

“…Time-resolved detection using pulsed lasers and time-gated measurements can also be used to address the signal-to-background problem in biological samples. This is done by limiting light detection to non-resonant interaction phenomena with relaxation time scales occurring on sub-nanosecond timescales, thereby excluding contributions from resonant phenomena such as fluorescence 36 …”

Development of a near-infrared Raman spectroscopy setup compatible with fluorescence-guided surgery

“…To overcome the interference of intense fluorescence emission, several approaches have been developed, including photo-bleaching, 18 confocal detection, 19 surface-enhanced Raman spectroscopy (SERS), 20 shifted-excitation Raman difference spectroscopy (SERDS), 21–23 polarization-resolved signal detection, 24 and time-resolved signal detection (time gating). 25–27 However, if feasible, the most straightforward way is to avoid the overlap of the RS excitation wavelength and fluorophore absorption wavelengths. 1 This approach was used by Puppels et al in 1993, where lymphocyte subpopulations were selected by means of fluorescence-activated cell sorting (FACS, 488 nm excitation), and Raman spectra were obtained with 660 nm laser excitation, thereby avoiding excitation of the fluorescent labels.…”