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A regression-based fusion algorithm has been used to merge hyperspectral Fourier transform infrared (FTIR) data with an H&E image of oral squamous cell carcinoma metastases in cervical lymphoid nodal tissue. This provides insight into the success of the ratio of FTIR absorbances at 1252 cm-1 and 1285 cm-1 in discriminating between these tissue types. The success is due to absorbances at these two wavenumbers being dominated by contributions from DNA and collagen, respectively. A pixel-by-pixel fit of the fused spectra to the FTIR spectra of collagen, DNA and cytokeratin reveals the contributions of these molecules to the tissue at high spatial resolution.

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A machine learning algorithm (MLA) has predicted the prognosis of oral potentially malignant lesions and discriminated between lymph node tissue and metastatic oral squamous cell carcinoma (OSCC). The MLA analyses metrics, which are ratios of Fourier transform infrared absorbances, and identifies key wavenumbers that can be associated with molecular biomarkers. The wider efficacy of the MLA is now shown in the more complex primary OSCC tumour setting, where it is able to identify seven types of tissue. Three epithelial and four non-epithelial tissue types were discriminated from each other with sensitivities between 82% and 96% and specificities between 90% and 99%. The wavenumbers involved in the five best discriminating metrics for each tissue type were tightly grouped, indicating that small changes in the spectral profiles of the different tissue types are important. The number of samples used in this study was small, but the information will provide a basis for further, larger investigations.

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A machine learning algorithm (MLA) has been applied to a Fourier transform infrared spectroscopy (FTIR) dataset previously analysed with a principal component analysis (PCA) linear discriminant analysis (LDA) model. This comparison has confirmed the robustness of FTIR as a prognostic tool for oral epithelial dysplasia (OED). The MLA is able to predict malignancy with a sensitivity of 84 ± 3% and a specificity of 79 ± 3%. It provides key wavenumbers that will be important for the development of devices that can be used for improved prognosis of OED.

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This work reports the first images obtained by combining an infrared aperture scanning near-field optical microscope (SNOM) with a quantum cascade laser (QCL). The future potential of this set-up is demonstrated by a preliminary study on an OE33 human oesophageal adenocarcinoma cell in which the cell is imaged at 1751 cm-1, 1651 cm-1, 1539 cm-1 and 1242 cm-1. In addition to the 1651 cm-1 image, three other images were acquired within the Amide I band (1689 cm-1, 1675 cm-1 and 1626 cm-1) chosen to correspond to secondary structures of proteins. The four images obtained within the Amide I band show distinct differences demonstrating the potential of this approach to reveal subtle changes in the chemical composition of a cell.

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Using a scanning near-field optical microscope coupled to an infrared free electron laser (SNOM-IR-FEL) in low-resolution transmission mode, we collected chemical data from whole cervical cells obtained from 5 pre-menopausal, non-pregnant women of reproductive age, and cytologically classified as normal or with different grades of cervical cell dyskaryosis. Imaging data are complemented by demography. All samples were collected before any treatment. Spectra were also collected using attenuated total reflection, Fourier-transform (ATR-FTIR) spectroscopy, to investigate the differences between the two techniques. Results of this pilot study suggests SNOM-IR-FEL may be able to distinguish cervical abnormalities based upon changes in the chemical profiles for each grade of dyskaryosis at designated wavelengths associated with DNA, Amide I/II, and lipids. The novel data sets are the first collected using SNOM-IR-FEL in transmission mode at the ALICE facility (UK), and obtained using whole cells as opposed to tissue sections, thus providing an 'intact' chemical profile. These data sets are suited to complementing future work on image analysis, and/or applying the newly developed algorithm to other datasets collected using the SNOM-IR-FEL approach.

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Cervical cancer remains a major cause of morbidity and mortality among women, especially in the developing world. Increased synthesis of proteins, lipids and nucleic acids is a pre-condition for the rapid proliferation of cancer cells. We show that scanning near-field optical microscopy, in combination with an infrared free electron laser (SNOM-IR-FEL), is able to distinguish between normal and squamous low-grade and high-grade dyskaryosis, and between normal and mixed squamous/glandular pre-invasive and adenocarcinoma cervical lesions, at designated wavelengths associated with DNA, Amide I/II and lipids. These findings evidence the promise of the SNOM-IR-FEL technique in obtaining chemical information relevant to the detection of cervical cell abnormalities and cancer diagnosis at spatial resolutions below the diffraction limit (≥0.2 µm). We compare these results with analyses following attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy; although this latter approach has been demonstrated to detect underlying cervical atypia missed by conventional cytology, it is limited by a spatial resolution of ~3 µm to 30 µm due to the optical diffraction limit.

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