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Federico Vernuccio

Publications and source records attributed to Federico Vernuccio.

4 recordsLinked to original sources

Tunable Dynamic Speckle Generation for Random Illumination Microscopy

Speckled illumination enhances widefield fluorescence microscopy by enabling optical sectioning and super resolution. In random illumination microscopy, sequences of speckled illumination patterns are used to excite fluorescent samples and images are reconstructed based on a statistical analysis of the intensity fluctuations. Although random illumination microscopy has been shown to give excellent performance, its widespread implementation is hindered by the high cost and complexity of the generation of suitable speckled illumination patterns, which is achieved using digital micro-mirror devices or spatial light modulators. Here, we present a zwitterion-doped liquid crystal (LC) device capable of generating independent, high-contrast speckle patterns with a tunable decorrelation time in the 0.1 s to 0.1 ms range under visible laser illumination. This LC-based dynamic speckle generator is applied to widefield random illumination fluorescence microscopy of tissue and cell samples, where it enables optical sectioning with a 2 micron axial resolution, and a 1.5-fold improvement in lateral spatial resolution. Owing to its low cost and simplicity, this LC speckle generator offers an attractive alternative to digital micro-mirror and spatial light modulator devices for implementing widefield random illumination microscopy.

physics.optics

Widefield two-photon random illumination microscopy (2P-RIM)

Biological and biomedical samples are routinely examined using focused two-photon (2P) fluorescence microscopy due to its intrinsic axial sectioning and reduced out-of-focus bleaching. However, 2P imaging often requires excitation intensities that can damage samples through ionization and radical formation. Additionally, the lateral resolution of 2P microscopy is lower compared to linear one-photon (1P) fluorescence microscopy. Widefield 2P microscopy, using cameras, holds promise for reducing photo-toxicity while maintaining high image acquisition rates. Widefield imaging trades the high power and short integration times of sequential single point scanning for the low power and extended integration times of parallel detection across millions of pixels. However, generating effective axial sectioning over arbitrarily large fields of view (FOVs) has remained a challenge. In this work, we introduce 2P Random Illumination Microscopy (2P-RIM), an easy-to-implement 2P widefield technique, that achieves low photo-damage, fast imaging, micrometric axial sectioning, and enhanced lateral resolution for arbitrarily large FOVs. By using widefield speckled illuminations in conjunction with an image standard deviation matching algorithm, 2P-RIM demonstrated multicolor imaging over FOVs greater than 200 um, lateral resolution 220 nm, axial sectioning 2 um, and peak excitation powers about 10 times lower than those used in focused laser scanning microscopy.

physics.optics

Phonon dephasing times determined with time-delayed, broadband CARS

Coherent Raman scattering techniques as coherent anti-Stokes Raman scattering (CARS), offer significant advantages in terms of pixel dwell times and speed as compared to spontaneous Raman scattering for investigations of crystalline materials. However, the spectral information in CARS is often hampered by the presence of a non-resonant contribution to the scattering process that shifts and distorts the Raman peaks. In this work, we apply a method to obtain non-resonant background-free spectra based on time-delayed, broadband CARS (TD-BCARS) using an intra-pulse excitation scheme. In particular, this method can measure the phononic dephasing times across the full phonon spectrum at once. We test the methodology on amorphous SiO2 (glass), which is used to characterize the setup-specific and material-independent response times, and then apply TD-BCARS to the analysis of single crystals of diamond and ferroelectrics of potassium titanyl phosphate (KTP) and potassium titanyl arsenate (KTA). For diamond, we determine a dephasing time of t = 7.81 ps for the single sp3 peak.

cond-mat.mtrl-sci

Comparing Transmission- and Epi-BCARS: A Transnational Round Robin on Solid State Materials

Broadband coherent anti-Stokes Raman scattering (BCARS) is an advanced Raman spectroscopy method that combines the spectral sensitivity of spontaneous Raman scattering (SR) with the increased signal intensity of single-frequency coherent Raman techniques. These two features make BCARS particularly suitable for ultra-fast imaging of heterogeneous samples, as already shown in biomedicine. Recent studies demonstrated that BCARS also shows exceptional spectroscopic capabilities when inspecting crystalline materials like lithium niobate and lithium tantalate, and can be used for fast imaging of ferroelectric domain walls. These results strongly suggest the extension of BCARS towards new imaging applications like mapping defects, strain, or dopant levels, similar to standard SR imaging. Despite these advantages, BCARS suffers from a spurious and chemically unspecific non-resonant background (NRB) that distorts and shifts the Raman peaks. Post-processing numerical algorithms are then used to remove the NRB and to obtain spectra comparable to SR results. Here, we show the reproducibility of BCARS by conducting an internal Round Robin with two different BCARS experimental setups, comparing the results on different crystalline materials of increasing structural complexity: diamond, 6H-SiC, KDP, and KTP. First, we compare the detected and phase-retrieved signals, the setup-specific NRB-removal steps, and the mode assignment. Subsequently, we demonstrate the versatility of BCARS by showcasing how the selection of pump wavelength, pulse width, and detection geometry can be tailored to suit the specific objectives of the experiment. Finally, we compare and optimize measurement parameters for the high-speed, hyperspectral imaging of ferroelectric domain walls in lithium niobate.

cond-mat.mtrl-sci