Biology subjects

Debarre, D.

Publications and source records attributed to Debarre, D..

8 recordsLinked to original sources

bioRxiv2026

Influence of Non-Specific Surface Adhesion on the Shape and Microrheology of Red Blood Cells

Poly-L-Lysine (PLL) mediates the non-specific adhesion of cells and is commonly used in Atomic Force Microscopy (AFM) measurements, to ensure that cells remain attached to the substrate. However, it is acknowledged that adhesion affects the measured mechanical properties, in particular in the case Red Blood Cells (RBCs). This results in a wide range of Youngs modulus E reported in the literature. The present study aims at providing a systematic approach to the impact of non-specific adhesion on the rheology of RBCs. It provides a correlation between the topography profile of adherent RBCs and their rheology, from weak (cPLL = 10-3 mg/mL) to strong-adhesion (cPLL = 100 mg/mL) regimes. Using RICM and AFM, we find that there is a continuum of RBC shapes promoted by adhesion, from concave to dome-shaped, as predicted by the theory of vesicle adhesion. Their elastic properties discriminate them into two populations depending on adhesion strength, where stiffer RBCs (E {gtrsim} 100 Pa) correlate with dome-shaped cells. These findings are supported by rheology measurements of the dynamic complex shear modulus G*(f): while the storage modulus increases with cell-substrate adhesion, reflective of an increased membrane shear modulus, the loss modulus remains unchanged. Finally, further analysis inspired by membrane theory shows that different deformation modes may be triggered during indentation of either weakly or strongly adhering RBCs, illustrating the limits of the Hertz model.

biophysics↗

bioRxiv2026

Quantifying optical sectioning in reflection microscopy with patterned illumination

Sectioning techniques based on patterned illumination have been widely used to obtain well-contrasted images of thick samples using widefield imaging setups. While their application to fluorescence microscopy has been extensively demonstrated and studied, their application to reflection imaging is scarcer and their performance has only been partly characterized. In this paper, we study numerically and analytically two such sectioning techniques, line confocal (LC) and structured illumination microscopy (SIM), in the context of their application to coherent reflection imaging. We derive approximate analytical equations to relate the performance of sectioning to the optical setup parameters, allowing straightforward understanding of their influence on the image intensity and depth of focus, and we systematically compare our predictions with experimental data. Finally, we quantify the precision and accuracy of each method in typical practical cases, providing guidelines to choose the most appropriate (LC, SIM, or a simple background subtraction on a widefield image) for the sample under study. We illustrate optical sectioning in the particular case of reflection interference contrast (RIC) microscopy, an imaging technique widely used in soft matter and biophysics studies to monitor object-surface interactions, or quantify surface functionalization. Our derivation, however, should also prove useful for other reflection methods such as optical coherence tomography (OCT) or flood illumination ophtalmoscopy.

biophysics↗

bioRxiv2025

Bacterial exploration of solid/liquid interfaces: developing platforms to control the physico-chemical microenvironment

Bacterial long-term contamination of surfaces is a promiscuous phenomenon often linked to harmful processes. Early bacterial exploration of interfaces, governed by adhesion and individual motility, is a known determinant of the subsequent development and persistence of bacterial colonies. However, the mechanisms by which bacteria integrate various environmental signals at these interfaces and modulate their behavior in response remain poorly understood. Here we present methods for designing precisely controlled microenvironments that enable the manipulation of both physical and chemical properties of solid-liquid interfaces, and also permit in situ monitoring of bacteria at these interfaces within microfluidic flow cells. Our aim is to provide an innovative toolbox for the interdisciplinary research community focused on elucidating the complex processes underlying bacterial surface exploration. We illustrate its use here by examining the surface motility of the pathogen Pseudomonas aeruginosa.

biophysics↗

bioRxiv2024

Optical sectioning for reflection interference microscopy

Reflection Interference Contrast Microscopy (RICM, also known as interference reflection microscopy) and related techniques have become of wide interest to the biophysical, soft matter and biochemistry communities owing to their exquisite sensitivity for characterising thin films or individual nanoscopic objects adsorbed onto surfaces, or for monitoring cell-substrate interactions. Over the recent years, striking progresses have been made to improve the sensitivity and the quantitative analysis of RICM. Its use in more complex environments, with spurious reflections stemming from a variety of structures in the sample, remains however challenging. In this paper, we demonstrate two optical sectioning methods that effectively reduce such background and can be readily implemented in a conventional RICM setup: line confocal detection, and structured illumination microscopy. We characterise experimentally the benefits to image quality and demonstrate the use of the methods for quantitative imaging of complex biological and biomimetic samples: cellular membranes, thin organic films, surface biofunctionalization. We then discuss the benefits of each method and provide guidelines to arbitrate between sectioning and signal-to-noise ratio. Finally, we provide a detailed description of our experimental setup and a home-written image acquisition and processing software that should allow the interested reader to duplicate such a setup on a home-built or commercial microscope.

biophysics↗

bioRxiv2024

Uncovering the simple adhesive strategy of the Toxoplasma parasite for high-speed motility

Toxoplasma gondii is a protozoan parasite that has evolved a developmental morphotype called tachyzoite that navigates between cells and moves in and out of them in a wide repertoire of homeothermic hosts. Relying on a uniquely constant apicobasal bipolarity coupled to an actomyosin-driven retrograde surface flow, the tachyzoite has elaborated a molecular machinery to assemble transient anchoring contacts with the environment, which support the traction force required to power a typical helical gliding motility. Combining micropatterning with live, reflection interference contrast and expansion microscopies, we bring first nanoscale evidence that the tachyzoite needs to build only one apical anchoring contact with the substrate, thus spatially defining a minimal force transmission platform over which it can slide. We uncover that the apicobasal driven surface flow is set up in response to extracellular biochemical cues independent of adhesin release and tachyzoite-surface interactions, hence prior to motile activity. Furthermore, to identify the minimal adhesion requirements for helical gliding at the level of individual molecular species, we combine biochemical and biophysical quantitative assays based on tunable surface chemistry and quartz crystal microbalance with dissipation monitoring. These approaches uncover that glycosaminoglycan (GAG)-parasite interactions are sufficient to promote a productive contact for helical gliding and pave the way for the characterization of the structure and density of the molecules functionally engaged at this essential parasite-substrate mechanosensitive interface.

cell biology↗

bioRxiv2023

Competitive specific anchorage of molecules onto surfaces: quantitative control of grafting densities and contamination by free anchors

The formation of surfaces decorated with biomacromolecules such as proteins, glycans or nucleic acids with well-controlled orientations and densities is of critical importance for the design of in vitro models, e.g., synthetic cell membranes, and interaction assays. To this effect, ligand molecules are often functionalized with an anchor that specifically binds to a surface with a high density of binding sites, providing control over the presentation of the molecules. Here, we present a method to robustly and quantitatively control the surface density of one or several types of anchor-bearing molecules by tuning the relative concentrations of target molecules and free anchors in the incubation solution. We provide a theoretical background that relates incubation concentrations to the final surface density of the molecules of interest, and present effective guidelines towards optimizing incubation conditions for the quantitative control of surface densities. Focussing on the biotin anchor, a commonly used anchor for interaction studies, as a salient example, we experimentally demonstrate surface density control over a wide range of densities and target molecule sizes. Conversely, we show how the method can be adapted to quality control the purity of end-grafted biopolymers such as biotinylated glycosaminoglycans by quantifying the amount of residual free biotin reactant in the sample solution.

biophysics↗

bioRxiv2022

A quantitative method for sizing hyaluronan and other glycosaminoglycans on surfaces

Hyaluronan (HA) is a major component of peri- and extra-cellular matrices and plays important roles in many biological processes such as cell adhesion, proliferation and migration. The abundance, size distribution and presentation of HA dictate its biological effects and are also useful indicators of pathologies and disease progression. Methods to assess the molecular mass of free-floating HA and other glycosaminoglycans (GAGs) are well established. In many biological and technological settings, however, GAGs are displayed on surfaces, and methods to obtain the size of surface-attached GAGs are lacking. Here, we present a method to size HA that is end-attached to surfaces. The method is based on the quartz crystal microbalance with dissipation monitoring (QCM-D) and exploits that the softness and thickness of films of grafted HA increase with HA size. These two quantities are sensitively reflected by the ratio of the dissipation shift ({Delta}D) and the negative frequency shift (-{Delta}f) measured by QCM-D upon the formation of HA films. Using a series of size-defined HA preparations, ranging in size from [~]2 kDa tetrasaccharides to [~]1 MDa polysaccharides, we establish a monotonic yet non-linear standard curve of the {Delta}D/-{Delta}f ratio as a function of HA size, which reflects the distinct conformations adopted by grafted HA chains depending on their size and surface coverage. We demonstrate that the standard curve can be used to determine the mean size of HA, as well as other GAGs, such as chondroitin sulfate and heparan sulfate, of preparations of previously unknown size in the range from 1 to 500 kDa, with a resolution of better than 10%. For polydisperse samples, our analysis shows that the process of surface-grafting preferentially selects smaller GAG chains, and thus reduces the average size of GAGs that are immobilised on surfaces comparative to the original solution sample. Our results establish a quantitative method to size HA and other GAGs grafted on surfaces, and also highlight the importance of sizing GAGs directly on surfaces. The method should be useful for the development and quality control of GAG-based surface coatings in a wide range of research areas, from molecular interaction analysis to biomaterials coatings.

biophysics↗

bioRxiv2022

Substrate stiffness impacts early biofilm formation via a modulation of twitching motility

Surface-associated lifestyles dominate in the bacterial world. Large multicellular assemblies, called biofilms, are essential to the survival of bacteria in harsh environments, and are closely linked to antibiotic resistance in pathogenic strains. Biofilms stem from the surface colonization of a wide variety of substrates encountered by bacteria, from living tissues to inert materials. Here, we demonstrate experimentally that the promiscuous opportunistic pathogen Pseudomonas aeruginosa explores substrates differently based on their rigidity, leading to striking variations in biofilm structure, exopolysaccharides (EPS) distribution, strain mixing during co-colonization and phenotypic expression. Using simple kinetic models, we show that these phenotypes arise through a mechanical interaction between the elasticity of the substrate and the type IV pilus (T4P) machinery, that mediates the surface-based motility called twitching. Together, our findings reveal a new role for substrate softness in the spatial organization of bacteria in complex microenvironments, with far-reaching consequences on efficient biofilm formation.

biophysics↗