Log-Gaussian gamma processes for training Bayesian neural networks in Raman and CARS spectroscopies.

We propose an approach utilizing gamma-distributed random variables, coupled with log-Gaussian modeling, to generate synthetic datasets suitable for training neural networks. This addresses the challenge of limited real observations in various applications. We apply this methodology to both Raman and coherent anti-Stokes Raman scattering (CARS) spectra, using experimental spectra to estimate gamma process parameters. Parameter estimation is performed using Markov chain Monte Carlo methods, yielding a full Bayesian posterior distribution for the model which can be sampled for synthetic data generation. Additionally, we model the additive and multiplicative background functions for Raman and CARS with Gaussian processes. We train two Bayesian neural networks to estimate parameters of the gamma process which can then be used to estimate the underlying Raman spectrum and simultaneously provide uncertainty through the estimation of parameters of a probability distribution. We apply the trained Bayesian neural networks to experimental Raman spectra of phthalocyanine blue, aniline black, naphthol red, and red 264 pigments and also to experimental CARS spectra of adenosine phosphate, fructose, glucose, and sucrose. The results agree with deterministic point estimates for the underlying Raman and CARS spectral signatures.

Bayesian Quantification for Coherent Anti-Stokes Raman Scattering Spectroscopy.

We propose a Bayesian statistical model for analyzing coherent anti-Stokes Raman scattering (CARS) spectra. Our quantitative analysis includes statistical estimation of constituent line-shape parameters, the underlying Raman signal, the error-corrected CARS spectrum, and the measured CARS spectrum. As such, this work enables extensive uncertainty quantification in the context of CARS spectroscopy. Furthermore, we present an unsupervised method for improving spectral resolution of Raman-like spectra requiring little to no a priori information. Finally, the recently proposed wavelet prism method for correcting the experimental artifacts in CARS is enhanced by using interpolation techniques for wavelets. The method is validated using CARS spectra of adenosine mono-, di-, and triphosphate in water, as well as equimolar aqueous solutions of d-fructose, d-glucose, and their disaccharide combination sucrose.

In vivo multiplex molecular imaging of vascular inflammation using surface-enhanced Raman spectroscopy.

Vascular immune-inflammatory responses play a crucial role in the progression and outcome of atherosclerosis. The ability to assess localized inflammation through detection of specific vascular inflammatory biomarkers would significantly improve cardiovascular risk assessment and management; however, no multi-parameter molecular imaging technologies have been established to date. Here, we report the targeted in vivo imaging of multiple vascular biomarkers using antibody-functionalized nanoparticles and surface-enhanced Raman scattering (SERS). Methods: A series of antibody-functionalized gold nanoprobes (BFNP) were designed containing unique Raman signals in order to detect intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and P-selectin using SERS. Results: SERS and BFNP were utilized to detect, discriminate and quantify ICAM-1, VCAM-1 and P-selectin in vitro on human endothelial cells and ex vivo in human coronary arteries. Ultimately, non-invasive multiplex imaging of adhesion molecules in a humanized mouse model was demonstrated in vivo following intravenous injection of the nanoprobes. Conclusion: This study demonstrates that multiplexed SERS-based molecular imaging can indicate the status of vascular inflammation in vivo and gives promise for SERS as a clinical imaging technique for cardiovascular disease in the future.

Automated replication of cone beam CT-guided treatments in the Pinnacle(3) treatment planning system for adaptive radiotherapy.

INTRODUCTION: Time-consuming manual methods have been required to register cone-beam computed tomography (CBCT) images with plans in the Pinnacle(3) treatment planning system in order to replicate delivered treatments for adaptive radiotherapy. These methods rely on fiducial marker (FM) placement during CBCT acquisition or the image mid-point to localise the image isocentre. A quality assurance study was conducted to validate an automated CBCT-plan registration method utilising the Digital Imaging and Communications in Medicine (DICOM) Structure Set (RS) and Spatial Registration (RE) files created during online image-guided radiotherapy (IGRT). METHODS: CBCTs of a phantom were acquired with FMs and predetermined setup errors using various online IGRT workflows. The CBCTs, DICOM RS and RE files were imported into Pinnacle(3) plans of the phantom and the resulting automated CBCT-plan registrations were compared to existing manual methods. A clinical protocol for the automated method was subsequently developed and tested retrospectively using CBCTs and plans for six bladder patients. RESULTS: The automated CBCT-plan registration method was successfully applied to thirty-four phantom CBCT images acquired with an online 0 mm action level workflow. Ten CBCTs acquired with other IGRT workflows required manual workarounds. This was addressed during the development and testing of the clinical protocol using twenty-eight patient CBCTs. The automated CBCT-plan registrations were instantaneous, replicating delivered treatments in Pinnacle(3) with errors of ±0.5 mm. These errors were comparable to mid-point-dependant manual registrations but superior to FM-dependant manual registrations. CONCLUSION: The automated CBCT-plan registration method quickly and reliably replicates delivered treatments in Pinnacle(3) for adaptive radiotherapy.

Preferential Attachment of Specific Fluorescent Dyes and Dye Labeled DNA Sequences in a Surface Enhanced Raman Scattering Multiplex.

A significant advantage of using surface enhanced Raman scattering (SERS) for DNA detection is the capability to detect multiple analytes simultaneously within the one sample. However, as the analytes approach the metallic surface required for SERS, they become more concentrated and previous studies have suggested that different dye labels will have different affinities for the metal surface. Here, the interaction of single stranded DNA labeled with either fluorescein (FAM) or tetramethylrhodamine (TAMRA) with a metal surface, using spermine induced aggregated silver nanoparticles as the SERS substrate, is investigated by analyzing the labels separately and in mixtures. Comparison studies were also undertaken using the dyes in their free isothiocyanate forms, fluorescein isothiocyanate (F-ITC) and tetramethylrhodamine isothiocyanate (TR-ITC). When the two dyes are premixed prior to the addition of nanoparticles, TAMRA exerts a strong masking effect over FAM due to a stronger affinity for the metal surface. When parameters such as order of analyte addition, analysis time, and analyte concentration are investigated, the masking effect of TAMRA is still observed but the extent changes depending on the experimental parameters. By using bootstrap estimation of changes in SERS peak intensity, a greater insight has been achieved into the surface affinity of the two dyes as well as how they interact with each other. It has been shown that the order of addition of the analytes is important and that specific dye related interactions occur, which could greatly affect the observed SERS spectra. SERS has been used successfully for the simultaneous detection of several analytes; however, this work has highlighted the significant factors that must be taken into consideration when planning a multiple analyte assay.

The recycling endosome protein Rab17 regulates melanocytic filopodia formation and melanosome trafficking.

Rab GTPases including Rab27a, Rab38 and Rab32 function in melanosome maturation or trafficking in melanocytes. A screen to identify additional Rabs involved in these processes revealed the localization of GFP-Rab17 on recycling endosomes (REs) and melanosomes in melanocytic cells. Rab17 mRNA expression is regulated by microphthalmia transcription factor (MITF), a characteristic of known pigmentation genes. Rab17 siRNA knockdown in melanoma cells quantitatively increased melanosome concentration at the cell periphery. Rab17 knockdown did not inhibit melanosome maturation nor movement, but it caused accumulation of melanin inside cells. Double knockdown of Rab17 and Rab27a indicated that Rab17 acts on melanosomes downstream of Rab27a. Filopodia are known to play a role in melanosome transfer, and in Rab17 knockdown cells filopodia formation was inhibited. Furthermore, we show that stimulation of melanoma cells with α-melanocyte-stimulating hormone induces filopodia formation, supporting a role for filopodia in melanosome release. Cell stimulation also caused redistribution of REs to the periphery, and knockdown of additional RE-associated Rabs 11a and 11b produced a similar accumulation of melanosomes and melanin to that seen after loss of Rab17. Our findings reveal new functions for RE and Rab17 in pigmentation through a distal step in the process of melanosome release via filopodia.