SignificancePancreatic surgery is a highly demanding and routinely applied procedure for the treatment of several pancreatic lesions. The outcome of patients with malignant entities crucially depends on the margin resection status of the tumor. Frozen section analysis for intraoperative evaluation of tissue is still time consuming and laborious.AimWe describe the application of fiber-based attenuated total reflection infrared (ATR IR) spectroscopy for label-free discrimination of normal pancreatic, tumorous, and pancreatitis tissue. A pilot study for the intraoperative application was performed.ApproachThe method was applied for unprocessed freshly resected tissue samples of 58 patients, and a classification model for differentiating between the distinct tissue classes was established.ResultsThe developed three-class classification model for tissue spectra allows for the delineation of tumors from normal and pancreatitis tissues using a probability score for class assignment. Subsequently, the method was translated into intraoperative application. Fiber optic ATR IR spectra were obtained from freshly resected pancreatic tissue directly in the operating room.ConclusionOur study shows the possibility of applying fiber-based ATR IR spectroscopy in combination with a supervised classification model for rapid pancreatic tissue identification with a high potential for transfer into intraoperative surgical diagnostics.
Pancreatic surgery is a highly demanding and routinely applied procedure for the treatment of several pancreatic lesions. The outcome of patients with malignant entities crucially depends on the margin resection status of the tumor. In this study we describe the application of fiber-based attenuated total reflection infrared (ATR IR) spectroscopy for label-free discrimination of normal pancreatic, tumorous and pancreatitis tissue. The method was applied for the unprocessed freshly resected tissue samples of 40 patients, and a classification model for differentiating between the distinct tissue classes was established. The developed three-class classification model for tissue spectra allows the delineation of tumors from normal and pancreatitis tissues. The classification algorithm provides probability values for each sample to be assigned to normal, tumor or pancreatitis classes. The established probability values were transferred to a Red-Green-Blue (RGB) color plot. Subsequently, the method was translated into intraoperative application. Fiber optic ATR IR spectra were obtained from freshly resected pancreatic tissue directly in the operating room. The spectroscopic findings could subsequently be confirmed by the histology gold standard. This study shows the possibility of applying fiber-based ATR IR spectroscopy in combination with a supervised classification model for rapid pancreatic tissue identification with a high potential for transfer into intraoperative surgical diagnostics.
The crucial goal of kidney-sparing surgical resection of a malignant tumor is complete removal of the cancerous tissue. The exact border between the cancerous and normal tissues is not always possible to identify by naked eye, therefore, a supplementary intraoperative diagnosis is needed. Unfortunately, intraoperative pathology methods used nowadays are time consuming and of inadequate quality rendering not definitive diagnosis. It has recently been shown that ATR-FTIR spectroscopy can be used for fast discrimination between cancerous and normal kidney tissues by analyzing the collected spectra of the tissue touch imprint smears. Most prominent differences are obtained in the wavenumber region from 950 cm-1 to 1250 cm-1, where the spectral bands due to the molecular vibrations of glycogen arise in the spectra of cancerous tissue smears. Such method of detection of cancerous tissue is limited by requirement to transfer the suspected tissue from the body to the FTIR instrument and stamp it on an ATR crystal of the spectrometer. We propose a spectroscopic tool which exploits the same principle of detection of cancerous cells as mentioned above, but does not require the tissue to be transferred from the body to the spectrometer. The portable spectrometer used in this design is equipped with fiber ATR probe and a sensitive liquid nitrogen cooled MCT detector. The design of the fiber probe allows the ATR tip to be changed easily in order to use only new sterilized tips for each measurement point of the tissue. It also enables sampling multiple areas of the suspected tissue with high lateral resolution which, in turn, increases accuracy with which the marginal regions between normal and cancerous tissues can be identified. Due to the loss of optical signal in the fiber probe the spectra have lower signal-to-noise ratio than in the case of standard ATR sampling setup. However, software for the spectral analysis used with the fiber probe design is still able to distinguish between cancerous and normal tissues with high accuracy.
Surface enhanced Raman scattering (SERS) spectroscopy is a useful method for detection of trace amounts of molecules. It has already been successfully implemented for detection of explosives, food additives, biomarkers in blood or urine, etc. In the last decade, SERS spectroscopy was introduced into the field of health sciences and has been especially focused on early disease detection. In the recent years, application of SERS spectroscopy for detection of various types of human cancerous tissues emerged. Furthermore, SERS spectroscopy of extracellular fluid shows great potential for the differentiation of normal and cancerous tissues; however, due to high variety of molecules present in such biological samples, the experimental spectrum is a combination of many different overlapping vibrational spectral bands. Thus, precise assignment of these bands to the corresponding molecular vibrations is a difficult task. In most cases, researchers try to avoid this task satisfying just with tentative assignment. In this study, low temperature SERS measurements of extracellular fluid of cancerous and healthy kidney tissue samples were carried out in order to get a deeper understanding of the nature of vibrational spectral bands present in the experimental spectrum. The SERS spectra were measured in temperature range from 300 K down to 100 K. SERS method was implemented using silver nanoparticle colloidal solution. The results of the low temperature SERS experiment were analysed and compared with the results of theoretical calculations. The analysis showed that the SERS spectrum of extracellular fluid of kidney tissue is highly influenced by the vibrational bands of adenine and Lcystine molecules.
Determination of cancerous and normal kidney tissues during partial, simple or radical nephrectomy surgery was performed by using differences in the IR absorption spectra of extracellular fluid taken from the corresponding tissue areas. The samples were prepared by stamping of the kidney tissue on ATR diamond crystal. The spectral measurements were performed directly in the OR during surgery for 58 patients. It was found that intensities of characteristic spectral bands of glycogen (880-1200 cm-1) in extracellular fluid are sensitive to the type of the tissue and can be used as spectral markers of tumours. Characteristic spectral band of lactic acid (1730 cm-1) - product of the anaerobic glycolysis, taking place in the cancer cells is not suitable for use as a spectral marker of cancerous tissue, since it overlaps with the band of carbonyl stretch in phospholipids and fatty acids. Results of hierarchical cluster analysis of the spectra show that the spectra of healthy and tumour tissue films can be reliably separated into two groups. On the other hand, possibility to differentiate between tumours of different types and grades remains in question. While the fluid from highly malignant G3 tumour tissue contains highly pronounced glycogen spectral bands and can be well separated from benign and G1 tumours by principal component analysis, the variations between spectra from sample to sample prevent from obtaining conclusive results about the grouping between different tumour types and grades. The proposed method is instant and can be used in situ and even in vivo.
Raman spectroscopy is known to provide information about the quality of the single walled carbon nanotubes (SWCNT).
The information is based on the intensity ratio of D and G spectral modes and the frequency of RBM modes. However
due to resonance nature of Raman spectrum of the nanotubes this method is not suitable to detect functionalization of the
nanotubes. Surface enhanced Raman spectroscopy (SERS) is known to enhance the Raman bands up to fourteen orders
of magnitude. Preferable adsorption sites for small silver nanoparticles are expected to be the functional groups of
SWCNT; therefore SERS technique allows detecting small amounts of functional groups despite strong resonance
Raman from backbone of SWCNT. In this study functionalized nanotubes were dispersed in silver colloid and dried on
the standard silver plate for Raman measurements. Spectra of SWCNT without colloid in the spectral range between 50
and 1800 cm-1 exhibit only four main spectral features: G, D, and RBM modes between 200 and 400 cm-1. Spectra of
SWCNT with the colloid exhibit several additional spectral bands which do not belong to the colloid. These bands
attributed to vibrations of C-O, C-C and O-H from the functional groups and the carbon atom of the SWCNT attached to
the corresponding group. The bands associated with the vibrations involving O atom is an indication that silver
nanoparticles interact with the functional group attached to SWCNT.
We present a novel approach to the detection of cancerous kidney tissue areas by measuring vibrational spectra (IR absorption or SERS) of intercellular fluid taken from the tissue. The method is based on spectral analysis of cancerous and normal tissue areas in order to find specific spectral markers. The samples were prepared by sliding the kidney tissue over a substrate - surface of diamond ATR crystal in case of IR absorption or calcium fluoride optical window in case of SERS. For producing the SERS signal the dried fluid film was covered by silver nanoparticle colloidal solution. In order to suppress fluorescence background the measurements were performed in the NIR spectral region with the excitation wavelength of 1064 nm. The most significant spectral differences – spectral markers - were found in the region between 400 and 1800 cm-1, where spectral bands related to various vibrations of fatty acids, glycolipids and carbohydrates are located. Spectral markers in the IR and SERS spectra are different and the methods can complement each other. Both of them have potential to be used directly during surgery. Additionally, IR absorption spectroscopy in ATR mode can be combined with waveguide probe what makes this method usable in vivo.
We have used infrared microspectroscopy for chemical analysis of urinary sediments. We showed that Mie scattering from urinary sediments as small as 10-100 μm is influencing the spectra and the influence can be
suppressed and quality of the spectra can be improved by applying RMieS-EMSC procedure.
Fourier transform infrared (FTIR) spectroscopic imaging has been used to probe the biochemical composition of human renal tumor tissue and adjacent normal tissue. Freshly resected renal tumor tissue from surgery was prepared as a thin cryosection and examined by FTIR spectroscopic imaging. Tissue types could be discriminated by utilizing a combination of fuzzy k-means cluster analysis and a supervised classification algorithm based on a linear discriminant analysis. The spectral classification is compared and contrasted with the histological stained image. It is further shown that renal tumor cells have spread in adjacent normal tissue. This study demonstrates that FTIR spectroscopic imaging can potentially serve as a fast and objective approach for discrimination of renal tumor tissue from normal tissue and even in the detection of tumor infiltration in adjacent tissue.
Infrared spectroscopic imaging of cancerous kidney tissue was performed by means of FTIR microscopy. The spectra of
thin tissue cryosections were collected with 64x64 MCT FPA detector and imaging area was increased up to 5.4×5.4 mm
by mapping by means of PC controlled x,y stage. Chemical images of the samples were constructed using statistical
treatment of the raw spectra. Several unsupervised and supervised statistical methods were used. The imaging results are
compared with results of the standard histopathological analysis. It was concluded that application of method of cluster
analysis ensures the best contrast of the images. It was found that border between cancerous and normal tissues visible in
the infrared spectroscopic image corresponds with the border visible in histopathological image. Closer examination of
the infrared spectroscopic image reveals that small domains of cancerous cells are found beyond the border in areas
distant from the border up to 3 mm. Such domains are not visible in the histopathological images. The smallest domains
found in the infrared images are approx. 60 μm.
Polarization Modulation Infrared Reflection Absorption Spectroscopy (PM-IRRAS) is a very sensitive imaging
technique for the characterization of molecular films. In order to achieve a spatial resolution close to the diffraction limit
a very small pinhole which acts as a point-source has to be used. However, such a small pinhole, the typical diameter
would be app. 100 μm, may reduces dramatically the intensity of the infrared beam. Using a common FTIR spectrometer
the spatial resolution is mainly limited by the brilliance of the globar infrared source. Therefore, an improvement in
lateral resolution requires a more brilliant light source.
The free electron laser (FEL) is such a high brilliant infrared source. The combination of the FEL with the PM-IRRAS
imaging system is a new approach to capture spectroscopic images with an excellent spatial resolution close to the
diffraction limit. PM-IRRAS images of a self assembly monolayer of phosphonic acid molecules onto a microstructures
gold / aluminum oxide surface where characterized. The spectroscopic image exhibits a spatial resolution of app. 5 μm.
An evaluation of characteristic absorbance bands of the phosphate group reveals that phosphonic acid molecules bound
with a high degree of orientation but differently at the gold and aluminum oxide surfaces. However, the spectroscopic
image reveals also several domains of disordering across the surface. Such domains have a dimension of only few
micrometers and can be identified in a high resolved PM-IRRAS image.
Access to the requested content is limited to institutions that have purchased or subscribe to SPIE eBooks.
You are receiving this notice because your organization may not have SPIE eBooks access.*
*Shibboleth/Open Athens users─please
sign in
to access your institution's subscriptions.
To obtain this item, you may purchase the complete book in print or electronic format on
SPIE.org.
INSTITUTIONAL Select your institution to access the SPIE Digital Library.
PERSONAL Sign in with your SPIE account to access your personal subscriptions or to use specific features such as save to my library, sign up for alerts, save searches, etc.