Biology subjects

Gasparin, F.

Publications and source records attributed to Gasparin, F..

4 recordsLinked to original sources

bioRxiv2026

Instantaneous Phase-Shifting Optothermal Microscopy for Live-cell Metabolic Monitoring

Cellular metabolic activities can be studied using label-free vibrational spectroscopic imaging to leverage the endogenous contrast of biomolecules. However, fast live-cell imaging over large fields-of-view remains challenging due to the need for raster scanning and despite advances in wide-field modalities, imaging rapid cellular activities across large cell populations remains challenging. Here, we introduce a mid-infrared optothermal microscopy method termed Instantaneous Phase-Shifting Optothermal Microscopy (IPSOM). IPSOM circumvents conventional mechanical phase-shifting methods, achieved single-frame imaging speed 588-fold increase compared to the sequential approach, under field-of-view of 300x350 {micro}m. For multi-wavelength hyperspectral imaging, IPSOM achieves an 8-fold speed improvement. IPSOM is used here to monitor lipid remodelling in adipocytes during lipolysis, demonstrating its potential for studying rapid cellular metabolism.

bioengineering↗

bioRxiv2026

Vibrational optoacoustic detection of lipid-membrane dynamics enables label-free imaging of cell membrane potential

Current technologies to monitor membrane potential are either highly invasive and perturb the integrity of the membrane, use labels that compromise biological behavior, or are limited in sensitivity and do not enable simultaneous monitoring of multiple cells and cell populations. Here, we present Mid-IR Assessment of Conformation in Lipids by Ensemble Sensing (MIRACLES) that, by detection of molecular vibration of lipid acyl chains under cell-membranes electric field dynamics, achieves highly sensitive label-free imaging of membrane potential dynamics in living cells. MIRACLES leverages lipid conformational changes within the plasma membrane as intrinsic indicator for cell membrane depolarization and hyperpolarization. As proof-of-concept, we apply MIRACLES to monitor membrane depolarization during glucose stimulated insulin secretion in {beta}-cells at single-cell level and achieve assessment of {beta}-cell functionality in real time. These results highlight the potential of mid-IR optoacoustic as a powerful tool for indirect, label-free potential assessment of cellular metabolic activities.

bioengineering↗

bioRxiv2024

Protein-structure-sensitive mid-infrared optoacoustic microscopy enables label-free assessment of drug therapy in myeloma cells

Conventional live-cell optical microscopy lacks sensitivity and specificity for label-free detection of intracellular protein-structure dynamics, such as conformational transition from -helix to {beta}-sheet. Detecting intermolecular {beta}-sheet formation, for instance, is important because it is a hallmark of misfolded proteins and aggresome formation--which are intrinsic indicators of cell apoptosis in myeloma therapy. Going beyond conventional optical microscopy, we introduce a single-cell imaging technology with label-free sensitivity to intracellular intermolecular {beta}-sheet formation in living cells. This unique ability was attained by exploiting the spectral specificity of the mid-infrared amide I region (1700 - 1600 cm-1) to protein structure and the positive-contrast nature of optoacoustic microscopy. By means of this technology, we were able to monitor the efficiency of proteasome inhibition in a myeloma cell line and--as a first demonstration towards clinical translation--in biopsied myeloma cells from patients. Achieving label-free monitoring of treatment at a single-cell level allows longitudinal assessment of response heterogeneity, which could provide crucial therapeutic information, such as patient-specific sensitivity to treatment, thus facilitating personalized medicine in myeloma therapy.

biophysics↗

bioRxiv2023

Phase-shifting mid-infrared optothermal microscopy for wide-field hyperspectral imaging of living cells

Fast live-cell hyperspectral imaging at large field-of-views (FOVs) and high cell confluency remains challenging in vibrational microscopy due to the need for point-by-point focal excitation scanning. Imaging at high cell confluency and large FOVs is important, respectively, for proper cell function and statistical significance of measurements. Here, we introduce phase-shifting mid-infrared optothermal microscopy (PSOM) which interprets molecular-vibrational information as the optical path difference (OPD) induced by mid-infrared absorption and is capable of taking snapshot vibrational images over broad mid-infrared excitation areas at high live-cell confluency. By means of phase-shifting, PSOM suppresses noise to a quarter of current optothermal microscopy modalities to allow capturing live-cell vibrational images at FOVs up to 50 times larger than state-of-the-art. Additionally, it reduces illumination power flux density (PFD) down to 5 orders of magnitude lower than conventional vibrational microscopy--thus, considerably decreasing the possibility of cell photodamage.

bioengineering↗