a-multimodal-wide-field-fourier-transform-raman-microscope

A Multimodal Wide-Field Fourier-Transform Raman Microscope

Raman microscopy is a powerful technique to determine the chemical composition of a sample. The main challenge of the state-of-the-art spontaneous Raman ...

Politecnico di Milano

IFN-CNR

We compared the effectiveness of three optical techniques based on fluorescence imaging and spectroscopy with indocyanine green (ICG) contrast agent to evaluate in vivo the disruption of the active vasculature induced by a vascular targeting agent. The blood perfusion of the MDA-MB-435 tumor model transplanted in nude mice was estimated from the signal of the contrast agent measured immediately after its systemic injection in mice. Optical measurements were performed using a fluorescence imaging setup and a fiber-based time correlated single photon counting (TCSPC) apparatus. This latter apparatus was used to measure the tumor fluorescence in transmittance geometry and the change in the basal optical absorption induced by the contrast agent, thus providing an alternative estimation of the blood content in the tumor. Mice were divided into four groups. Three groups were treated with different doses of the vascular disrupting agent ZD6126, the fourth group (control group) received the drug vehicle only. Optical measurements were carried out 3 h after pharmacologic treatment. After 24 h, mice were killed, tumors were excised and the extent of necrosis was evaluated with standard histologic analysis. On fluorescence imaging ICG emission from tumors of mice treated with ZD6126 significantly was lower compared with the emission from control mice. The histologic sections also showed a significantly higher amount of necrosis in tumors of treated mice. Both these findings, which correlate with each other, indicate an effective vascular shutdown induced by the drug. However, ICG fluorescence measured with the TCSPC apparatus in transmittance geometry and the estimate of the change in optical absorption did not allow a statistically significant differentiation between treated and control groups.

In vivo measurement of vascular modulation in experimental tumors using a fluorescent contrast agent.

In order to reduce both acquisition and reconstruction times, illumination and detection in fluorescence diffuse optical tomography (FDOT) have recently evolved from a point-based to a pattern-based approach. The use of structured illumination, offering the ability to project any pattern of light onto the object, associated with the compression of the acquired fluorescence images has paved the way for a new generation of fast reconstruction algorithms for FDOT. However, the choice of the most appropriate set of source patterns is still an open problem. Here, the use of typical source patterns is investigated on experimental data. Reconstructions of similar qualities are obtained for the different types of source patterns. We found that the performances of structured illumination are limited by the required positivity of the source patterns. To alleviate this problem, we introduce a novel method, namely the virtual source pattern method, which allows for considering any kind of patterns, e.g., with negative and complex intensities. This new method provides a significant increase of the contrast of the reconstruction and also a reduction of the reconstruction error, especially when virtual wavelet source patterns are considered.

A virtual source pattern method for fluorescence tomography with structured light.

Applications of time-resolved photoluminescence spectroscopy (TRPL) and fluorescence lifetime imaging (FLIM) to the analysis of cultural heritage are presented. Examples range from historic wall paintings and stone sculptures to 20th century iconic design objects. A detailed description of the instrumentation developed and employed for analysis in the laboratory or in situ is given. Both instruments rely on a pulsed laser source coupled to a gated detection system, but differ in the type of information they provide. Applications of FLIM to the analysis of model samples and for the in-situ monitoring of works of art range from the analysis of organic materials and pigments in wall paintings, the detection of trace organic substances on stone sculptures, to the mapping of luminescence in late 19th century paintings. TRPL and FLIM are employed as sensors for the detection of the degradation of design objects made in plastic. Applications and avenues for future research are suggested.

Time-resolved photoluminescence spectroscopy and imaging: new approaches to the analysis of cultural heritage and its degradation.

Calcium oscillations play a role in the regulation of the development of tip-growing plant cells. Using optical microscopy, calcium oscillations have been observed in a few systems (e.g. pollen tubes, fungal hyphae and algal rhizoids). High-resolution, non-phototoxic and rapid imaging methods are required to study the calcium oscillation in root hairs. We show that light sheet fluorescence microscopy is optimal to image growing root hairs of Arabidopsis thaliana and to follow their oscillatory tip-focused calcium gradient. We describe a protocol for performing live imaging of root hairs in seedlings expressing the cytosol-localized ratiometric calcium indicator Yellow Cameleon 3.6. Using this protocol, we measured the calcium gradient in a large number of root hairs. We characterized their calcium oscillations and correlated them with the rate of hair growth. The method was then used to screen the effect of auxin on the properties of the growing root hairs.

Light Sheet Fluorescence Microscopy Quantifies Calcium Oscillations in Root Hairs of Arabidopsis thaliana.

It is known that oil paintings containing zinc white are subject to rapid degradation. This is caused by the interaction between the active groups of binder and the metal ions of the pigment, which gives rise to the formation of new zinc complexes (metal soaps). Ongoing studies on zinc white paints have been limited to the chemical mechanisms that lead to the formation of zinc complexes. On the contrary, little is known of the photo-physical changes induced in the zinc oxide crystal structure following this interaction. Time-resolved photoluminescence spectroscopy has been applied to follow modifications in the luminescent zinc white pigment when mixed with binder. Significant changes in trap state photoluminescence emissions have been detected: the enhancement of a blue emission combined with a change of the decay kinetic of the well-known green emission. Complementary data from molecular analysis of paints using Fourier transform infrared spectroscopy confirms the formation of zinc carboxylates and corroborates the mechanism for zinc complexes formation. We support the hypothesis that zinc ions migrate into binder creating novel vacancies, affecting the photoluminescence intensity and lifetime properties of zinc oxide. Here, we further demonstrate the advantages of a time-resolved photoluminescence approach for studying defects in semiconductor pigments.

A Photoluminescence Study of the Changes Induced in the Zinc White Pigment by Formation of Zinc Complexes.

In conservation, science semiconductors occur as the constituent matter of the so-called semiconductor pigments, produced following the Industrial Revolution and extensively used by modern painters. With recent research highlighting the occurrence of various degradation phenomena in semiconductor paints, it is clear that their detection by conventional optical fluorescence imaging and microscopy is limited by the complexity of historical painting materials. Here, we illustrate and prove the capabilities of time-resolved photoluminescence (TRPL) microscopy, equipped with both spectral and lifetime sensitivity at timescales ranging from nanoseconds to hundreds of microseconds, for the analysis of cross-sections of paint layers made of luminescent semiconductor pigments. The method is sensitive to heterogeneities within micro-samples and provides valuable information for the interpretation of the nature of the emissions in samples. A case study is presented on micro samples from a painting by Henri Matisse and serves to demonstrate how TRPL can be used to identify the semiconductor pigments zinc white and cadmium yellow, and to inform future investigations of the degradation of a cadmium yellow paint.

Time-Resolved Photoluminescence Microscopy for the Analysis of Semiconductor-Based Paint Layers.

Accurate risk stratification for patients with non-dilated left ventricular cardiomyopathy (NDLVC) remains challenging due to lack of dedicated clinical trials. This post hoc analysis aims to delineate the arrhythmic risk and assess the incremental value of cardiac magnetic resonance (CMR) imaging in the CarDiac magnEtic Resonance for prophylactic Implantable-cardioVerter defibrillAtor ThErapy (DERIVATE) study cohort meeting the NDLVC diagnostic criteria.

CarDiac magnEtic Resonance for prophylactic Implantable cardioVerter defibrillAtor ThErapy in Non-Dilated Left Ventricular Cardiomyopathy: a sub-study from the DERIVATE registry.

Selection of the patients for implantable cardioverter defibrillator primary prevention therapy in non-ischaemic cardiomyopathy (NICM) needs to be improved. To evaluate the additional prognostic value of a new cardiac magnetic resonance (CMR) score based on late gadolinium enhancement (LGE) pattern distribution (DERIVATE Risk Score 2.0) when compared with previously published DERIVATE Risk Score 1.0, which is based solely on quantitative parameters, in a cohort of NICM patients enrolled in the DERIVATE registry.

Article	Year
In vivo measurement of vascular modulation in experimental tumors using a fluorescent contrast agent. Photochemistry and photobiology\| PubMed ID: 18422875	2008
A virtual source pattern method for fluorescence tomography with structured light. Physics in medicine and biology\| PubMed ID: 22641463	2012
Time-resolved photoluminescence spectroscopy and imaging: new approaches to the analysis of cultural heritage and its degradation. Sensors (Basel, Switzerland)\| PubMed ID: 24699285	2014
Light Sheet Fluorescence Microscopy Quantifies Calcium Oscillations in Root Hairs of Arabidopsis thaliana. Plant & cell physiology\| PubMed ID: 28379562	2017
A Photoluminescence Study of the Changes Induced in the Zinc White Pigment by Formation of Zinc Complexes. Materials (Basel, Switzerland)\| PubMed ID: 28772700	2017
Time-Resolved Photoluminescence Microscopy for the Analysis of Semiconductor-Based Paint Layers. Materials (Basel, Switzerland)\| PubMed ID: 29160862	2017
CarDiac magnEtic Resonance for prophylactic Implantable cardioVerter defibrillAtor ThErapy in Non-Dilated Left Ventricular Cardiomyopathy: a sub-study from the DERIVATE registry. European heart journal. Cardiovascular Imaging\| PubMed ID: 39899463	2025
Redefining the risk of major arrhythmic events in non-ischaemic cardiomyopathy: insights from the DERIVATE-NICM study. European heart journal. Cardiovascular Imaging\| PubMed ID: 40729425	2025

G. Valentini

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