Noninvasive simultaneous monitoring of pH and depth using surface-enhanced deep Raman spectroscopy

This is the final version. Available from Wiley via the DOI in this record. Here we demonstrate the simultaneous recovery of multiplexed physical information of surface-enhanced Raman scattering (SERS) nanoparticles (pH and depth) using deep Raman spectroscopy. As has been shown previously and in accordance with theory, inelastically scattered photons arising from spectral peaks that are suitably separated can exhibit different optical properties in the media through which they travel. These differences can impact the relative intensities of the Raman peaks as a function of the transmission path length; thereby, the depth of signal generation is inherently encoded in the spectra; assuming the target is clustered at a single depth or location, its depth can be readily determined. Moreover, Raman spectroscopy is very sensitive to chemistry of a sample, and changes in pH are observed not only as changes in peak intensity through relevant protonation and deprotonation but also as shifts in spectral features. Here, we show it is possible to precisely predict the depth (root-mean-square error [RMSE] 5 %) of SERS nanoparticles in scattering media (0.5% intralipid) while also being able to noninvasively monitor simultaneously the pH levels (RMSE ~0.2 pH units) of the media surrounding the nanoparticles. This is important as it demonstrates that nanoparticles can be used to report on multiple physical properties including their depth. This opens avenues for a range of new applications including the noninvasive diagnosis and localisation of cancer lesions in clinical environment in vivo.Engineering and Physical Sciences Research Council (EPSRC)Engineering and Physical Sciences Research Council (EPSRC)Engineering and Physical Sciences Research Council (EPSRC

Exploring the effect of laser excitation wavelength on signal recovery with deep tissue transmission Raman spectroscopy.

PublishedJOURNAL ARTICLEThe aim of this research was to find the optimal Raman excitation wavelength to attain the largest possible sensitivity in deep Raman spectroscopy of breast tissue. This involved careful consideration of factors such as tissue absorption, scattering, fluorescence and instrument response function. The study examined the tissue absorption profile combined with Raman scattering and detection sensitivity at seven different, laser excitation wavelengths in the near infrared region of the spectrum. Several key scenarios in regards to the sample position within the tissue were examined. The highest Raman band visibility over the background ratio in respect to biological tissue provides the necessary information for determining the optimum laser excitation wavelength for deep tissue analysis using transmission Raman spectroscopy, including detection of breast calcifications. For thick tissues with a mix of protein and fat, such as breast tissue, 790-810 nm is concluded to be the optimum excitation wavelength for deep Raman measurements.An EPSRC grant (EP/K020374/1) funded the work presented here

Sensitivity of Transmission Raman Spectroscopy Signals to Temperature of Biological Tissues

This is the final version of the article. Available from Springer Nature via the DOI in this record.Optical properties of biological tissues can be influenced by their temperature, thus affecting light transport inside the sample. This could potentially be exploited to deliver more photons inside large biological samples, when compared with experiments at room temperature, overcoming some of difficulties due to highly scattering nature of the tissue. Here we report a change in light transmitted inside biological tissue with temperature elevation from 20 to 40 °C, indicating a considerable enhancement of photons collected by the detector in transmission geometry. The measurement of Raman signals in porcine tissue samples, as large as 40 mm in thickness, indicates a considerable increase in signal ranging from 1.3 to 2 fold, subject to biological variability. The enhancements observed are ascribed to phase transitions of lipids in biological samples. This indicates that: 1) experiments performed on tissue at room temperature can lead to an underestimation of signals that would be obtained at depth in the body in vivo and 2) that experiments at room temperature could be modified to increase detection limits by elevating the temperature of the material of interest.The work was supported by a grant from the Engineering and Physical research council (EP/P012442/1)

Direct monitoring of light mediated hyperthermia induced within mammalian tissues using surface enhanced spatially offset Raman spectroscopy (T-SESORS)

This is the final version. Available on open access from Royal Society of Chemistry via the DOI in this recordData availability: the research data supporting this publication are provided within this paper.Here we demonstrate light mediated heating of nanoparticles confined deep inside mammalian tissue, whilst directly monitoring their temperature non-invasively using a form of deep Raman spectroscopy, T-SESORS. One of the main barriers to the introduction of photo-thermal therapies (PTT) has been recognised as the inability to directly monitor the local temperature deep within the tissue at the point of therapy. Here Au nanoparticles with a Raman reporter molecule (temperature reporters) are used in combination with Au nanoshells (heat mediators) to provide simultaneously heating under NIR illumination and direct spectroscopic monitoring of local temperature deep within mammalian tissues. The surface enhanced Raman signal was read out at the tissue surface using a transmission geometry in this example and the temperature of the tissue was ascertained from the anti-Stokes to Stokes Raman reporter. This approach opens the prospect of non-invasive hyperthermia treatments with direct temperature feedback from deep inside within tissue, where nanoparticles can be used to both provide localised heating and accurately monitor the local temperature.Engineering and Physical Sciences Research Council (EPSRC

Subsurface chemically specific measurement of pH levels in biological tissues using combined surface-enhanced and deep Raman

This is the final version. Available from the American Chemical Society via the DOI in this record. There is much interest in using nanosensors to monitor biologically relevant species such as glucose, or cellular pH, as these often become dysregulated in diseases such as cancer. This information is often inaccessible at depth in biological tissue, due to the highly scattering nature of tissue. Here we show that gold nanoparticles labeled with pH-sensitive reporter molecules can monitor pH at depth in biological tissues. This was achieved using deep Raman spectroscopy (spatially offset Raman and transmission Raman) in combination with surface-enhanced Raman spectroscopy, allowing chemical information to be retrieved significantly deeper than conventional Raman spectroscopy permits. Combining these approaches with chemometrics enabled pH changes to be monitored with an error of ±∼0.1 pH units noninvasively through 22 mm of soft tissue. This development opens the opportunity for the next generation of light-based medical diagnostic methods, such as monitoring of cancers, known to significantly alter pH levels.EPSR

Detection of age-related changes in tendon molecular composition by Raman spectroscopy—potential for rapid, non-invasive assessment of susceptibility to injury

The lack of clinical detection tools at the molecular level hinders our progression in preventing age-related tendon pathologies. Raman spectroscopy can rapidly and non-invasively detect tissue molecular compositions and has great potential for in vivo applications. In biological tissues, a highly fluorescent background masks the Raman spectral features and is usually removed during data processing, but including this background could help age differentiation since fluorescence level in tendons increases with age. Therefore, we conducted a stepwise analysis of fluorescence and Raman combined spectra for better understanding of the chemical differences between young and old tendons. Spectra were collected from random locations of vacuum-dried young and old equine tendon samples (superficial digital flexor tendon (SDFT) and deep digital flexor tendon (DDFT), total n = 15) under identical instrumental settings. The fluorescence-Raman spectra showed an increase in old tendons as expected. Normalising the fluorescence-Raman spectra further indicated a potential change in intra-tendinous fluorophores as tendon ages. After fluorescence removal, the pure Raman spectra demonstrated between-group differences in CH2 bending (1450 cm-1) and various ring-structure and carbohydrate-associated bands (1000-1100 cm-1), possibly relating to a decline in cellular numbers and an accumulation of advanced glycation end products in old tendons. These results demonstrated that Raman spectroscopy can successfully detect age-related tendon molecular differences

Studying the distribution of deep Raman spectroscopy signals using liquid tissue phantoms with varying optical properties

This is the final version of the article. Available from the publisher via the DOI in this record.In this study we employed large volume liquid tissue phantoms, consisting of a scattering agent (Intralipid), an absorption agent (Indian ink) and a synthesized calcification powder (calcium hydroxyapatite (HAP)) similar to that found in cancerous tissues (e.g. breast and prostate), to simulate human tissues. We studied experimentally the magnitude and origin of Raman signals in a transmission Raman geometry as a function of optical properties of the medium and the location of calcifications within the phantom. The goal was to inform the development of future noninvasive cancer screening applications in vivo. The results provide insight into light propagation and Raman scattering distribution in deep Raman measurements, exploring also the effect of the variation of relative absorbance of laser and Raman photons within the phantoms. Most notably when modeling breast and prostate tissues it follows that maximum signals is obtained from the front and back faces of the tissue with the central region contributing less to the measured spectrum.We thank the STFC BioMedical Network (STFC, STMA00012) and the University of Exeter for their financial support. An EPSRC grant [EP/K020374/1] partly funded the work presented here

Spectroscopic study of optically induced ultrafast electron dynamics in gold

Copyright © 2007 The American Physical SocietyUsing a supercontinuum pulse as a probe, we have measured the transient reflectivity spectra of a thin film of gold for different values of the pump-probe time delay. The wavelength lambda(x) at which the measured transient reflectivity changes sign has been found to depend upon the time delay, leading to bipolar time resolved signals. The time dependence of lambda(x) has been shown to be consistent with calculations that take into account the full dependence of the reflectivity upon the electron occupation number, and to contradict qualitatively a model in which the signal is assumed to be directly proportional to the occupation number. The shift of lambda(x) has been found to persist at time delays that are much longer than the time required for the electrons to thermalize. Therefore the bipolar reflectivity signals do not necessarily contain a contribution from nonthermalized electrons, as has been previously assumed

Fluorescence suppression using micro-scale spatially offset Raman spectroscopy

We present a new concept of fluorescence suppression in Raman microscopy based on micro-spatially offset Raman spectroscopy which is applicable to thin stratified turbid (diffusely scattering) matrices permitting the retrieval of the Raman signals of sublayers below intensely fluorescing turbid over-layers. The method is demonstrated to yield good quality Raman spectra with dramatically suppressed fluorescence backgrounds enabling the retrieval of Raman sublayer signals even in situations where conventional Raman microscopy spectra are fully overwhelmed by intense fluorescence. The concept performance was studied theoretically using Monte Carlo simulations indicating the potential of up to an order or two of magnitude suppression of overlayer fluorescence backgrounds relative to the Raman sublayer signals. The technique applicability was conceptually demonstrated on layered samples involving paints, polymers and stones yielding fluorescence suppression factors between 12 to above 430. The technique has potential applications in a number of analytical areas including cultural heritage, archaeology, polymers, food, pharmaceutical, biological, biomedical, forensics and catalytic sciences and quality control in manufacture