Biology subjects

Pasqualini, F. S.

Publications and source records attributed to Pasqualini, F. S..

3 recordsLinked to original sources

bioRxiv2025

LigHTS: Massively Parallel Biomimetic Photo-Functionalization for Imaging-Based Ultra-High-Throughput Screening

Imaging-based ultra-high-throughput screening (UHTS) in pharma and biotech still runs on 384/1536-well plates whose stiff, flat substrates limit biological fidelity and screening efficiency. Highly biomimetic organs-on-chips and organoids improve relevance but lack reproducibility and plate-scale throughput. Biomimetic hydrogel scaffolds can be produced at scale through photopolymerization, which yet uses focused optics to define micrometer-resolved geometries, constraining scalability. To address the technical challenge of truly scalable biomimetic substrates featuring anisotropies, this study presents LigHTS, an all-optical, in-well method that replaces focused with collimated illumination to photofabricate structured hydrogels in standard 384/1536-well plates. Adding food dye tartrazine to gelatin-methacrylate (GelMA) solutions increases hydrogel thickness sensitivity to UV dose by [~]10x, allowing uniform control of film thickness without lenses. Entire plates are functionalized in parallel with soft hydrogels ([~]1-10 kPa) whose thickness is tunable from 10 to 100 {micro}m. Simultaneously, simply interposing film photomasks encoding anisotropies enables orthogonal control of thickness and topography at UHTS throughput. Biological effect is demonstrated with mechanosensitive HT1080 cells, which display stiffness- and topography-dependent spreading and contact-guided migration on LigHTS-produced grooved substrates. Geometric uniformity across the plate (coefficient of variation <20%) meets HTS reproducibility standards, providing a readily available solution with enhanced biomimicry for imaging-based UTHS pipelines.

bioengineering↗

bioRxiv2025

BIOPOINT: A particle-based model for probing nuclear mechanics and cell-ECM interactions via experimentally derived parameters

Morphogenesis involves biochemical and biomechanical interactions across multiple spatial and temporal scales. Experimental studies alone struggle to resolve these dynamics, necessitating computational models. Among these, subcellular element modeling (SEM) has proven helpful in simulating cellular and tissue-scale emergent behaviors. However, traditional SEM frameworks lack explicit representations of nuclear mechanics and cell-extracellular matrix (ECM) interactions, limiting their ability to capture key biology. Here, we introduce BIOPOINT, a particle-based computational framework that extends SEM by incorporating (1) a deformable, multi-particle nucleus to simulate nuclear stress and strain distributions and (2) an explicit ECM representation using a structured array of static particles interacting via tunable adhesive potentials. To ensure biological relevance, we calibrated BIOPOINTs parameters against single-cell indentation experiments, overcoming prior limitations of ad hoc parameter selection in SEM. We validate BIOPOINT by comparing simulated cell behaviors to experimental observations in three key scenarios: (i) single-cell indentation, demonstrating agreement with force-time curves from atomic force microscopy (AFM) studies; (ii) cell spreading on ECM micropatterns, confirming that nuclear deformation follows ECM constraints; and (iii) nuclear deformation during confined migration, showing BIOPOINT predicts nuclear shape dynamics as cells traverse constrictions accurately. BIOPOINT provides a computational framework for simulating nuclear mechanics and cell-ECM interactions with experimentally derived parameters. By integrating experimental data with a particle-based approach, BIOPOINT offers a practical tool for studying cell behavior that can inform future morphogenetic studies in-vivo or in-vitro.

biophysics↗

bioRxiv2024

HYDRA: HYdrogel Dispensing with Robotic Automation for high throughput drug testing

Traditional plastic- and glass-based cell culture platforms lack physiological relevance which limits their predictive power in drug development. Organoids and organs-on-chip offer biomimicry in late preclinical stages but lack scalability for high-throughput screening (HTS) in earlier stages. In fact, even introducing the simplest critical biomimetic cue, thin hydrogel layers, into the small wells of HTS plates causes the formation of curved menisci that disrupt cell seeding and imaging. We introduce HYDRA (HYDrogels by Robotic liquid handling Automation), an automated method that enables consistent hydrogel fabrication in HTS plates. We used liquid handling robots to dispense hydrogel precursor solutions and avoid contact with the sidewalls to prevent meniscus formation. We used gelatin for its mild surfactant properties, which allow pinning of the droplets contact line, promoting uniform spread. Finally, we re-aspirated the excess liquid, leaving behind a hydrogel layer with controlled stiffness and thickness. We show compatibility with standard 96- and 384-well plates and integration with existing automated workflows, including quality control. We validated HYDRA in dose-response assays using anti-cancer drugs, genetically engineered epithelial cells, phase holography, and fluorescence microscopy. We believe that HYDRA provides a scalable, biomimetic, and therefore more predictive alternative for early- stage screening and in vitro drug testing.

bioengineering↗