Size and fluorescence measurements of individual droplets by flow cytometry

Fattaccioli, Jacques; Baudry, Jean; Émerard, Jean-Daniel; Bertrand, Emanuel; Goubault, Cécile; Henry, Nelly; Bibette, Jérôme

doi:10.1039/b814954b

Cited by 42 publications

(44 citation statements)

References 23 publications

(19 reference statements)

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“…Liquid and solid particles were characterized with a BD Accuri C6 cytometer (BD 22 Biosciences, New Jersey, USA) according to a method we developed in the past [36] for 23 quasi-monodisperse emulsions. In brief, the fluorescence intensity of the particles is 24…”

Section: Characterization Of Opsonization Degree Of the Particles 21mentioning

confidence: 99%

“…Carefully 14 functionalized with biologically-relevant adhesive molecules, they are able to interact with 15 cells in a specific manner [32] and can be used as model particles for cell adhesion modeling 16 [33][34][35]. In addition to giving access to a controlled range of biomolecules densities at the 17 surface, emulsion droplets have a liquid interface allowing proteins bound to it to be laterally 18 mobile [34][35][36], as in our case the IgGs. However, to our knowledge there is no report of any 19 observations at the scale of a single droplet interacting with a phagocyte, nor obvious 20 elements about the possible influence of the nature of the interface (liquid vs. solid) on the 21 uptake, despite its biophysical [37] and biological relevance [4,22,23].…”

Section: Introductionmentioning

confidence: 99%

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Phagocytosis of immunoglobulin-coated emulsion droplets

et al. 2015

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Phagocytosis by macrophages represents a fundamental process essential for both immunity and tissue homeostasis. The size of targets to be eliminated ranges from small particles as bacteria to large objects as cancerous or senescent cells. Most of our current quantitative knowledge on phagocytosis is based on the use of solid polymer microparticles as model targets that are well adapted to the study of phagocytosis mechanisms that do not involve any lateral mobility of the ligands, despite the relevance of this parameter in the immunological context. Herein we designed monodisperse, IgG-coated emulsion droplets that are efficiently and specifically internalized by macrophages through in-vitro FcγR-mediated phagocytosis. We show that, contrary to solid polymeric beads, droplet uptake is efficient even for low IgG densities, and is accompagnied by the clustering of the opsonins in the zone of contact with the macrophage during the adhesion step. Beyond the sole interest in the design of the material, our results suggest that lateral mobility of proteins at the interface of a target greatly enhances the phagocytic uptake.

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Section: Characterization Of Opsonization Degree Of the Particles 21mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Phagocytosis of immunoglobulin-coated emulsion droplets

et al. 2015

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“…On the other hand, the binding constant for the interaction between nitrilotriacetic acid (NTA) and histidine is much lower on the order of K b = 10 6 mol −1 6. Many important applications take advantage of the wide range of interaction strengths between ligand-receptor pairs to control the binding of different species 7–11. For example, in the field of targeted drug delivery, functionalization of polymer end groups can be used for specific targeting12–20 of cells.…”

Section: Introductionmentioning

confidence: 99%

Streptavidin−Biotin Binding in the Presence of a Polymer Spacer. A Theoretical Description

Ren

Carvajal²,

Shull³

et al. 2009

Langmuir

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The binding of streptavidin to biotin located at the terminal ends of poly(ethylene oxide) tethered to a planar surface is studied using molecular theory. The theoretical model is applied to mimic experiments (Langmuir 2008(Langmuir , 24, 2472 performed using drop-shape analysis to study receptorligand binding at the oil/water interface. Our theoretical predictions show very good agreements with the experimental results. Furthermore, the theory enables us to study the thermodynamic and structural behavior of the PEO-biotin+streptavidin layer. The interfacial structure, shown by the volume fraction profiles of bound proteins and polymers, indicates that the proteins form a thick layer supported by stretched polymers, where the distribution of bound proteins is greater than the thickness of the height of one layer of proteins. When the polymer spacer is composed of PEO (3000), a thick layer with multi-layers of proteins is formed, supported by the stretched polymer chains. It was found that thick multi-layers of proteins are formed when long spacers are present or at very high protein surface coverages on short spacers. This shows that the flexibility of the polymer spacer plays an important role in determining the structure of the bound proteins due to their ability to accommodate highly distorted conformations to optimize binding and protein interactions. Protein domains are predicted when the amount of bound proteins is small due to the existence of streptavidinstreptavidin attractive interactions. As the number of proteins is increased, the competition between attractive interactions and steric repulsions determines the stability and structure of the bound layer. The theory predicts that the competition between these two forces leads to a phase separation at higher protein concentrations. The point where this transition happens depends on both spacer length and protein surface coverage and is an important consideration for practical applications of these and other similar systems. If the goal is to maximize protein binding, it is favorable to be above the layer transition, as multiple layers can accommodate greater bound protein densities. On the other hand, if the goal is to use these bound proteins as a linker group to build more complex structures, such as when avidin or streptavidin serves as a linker between two biotinylated polymers or proteins, the optimum is to be below the layer transition such that all bound linker proteins are available for further binding. II. INTRODUCTIONLigand-receptor interactions are an important control mechanism in many biological systems. For example, the binding of extracellular matrix proteins to specific receptors on the cell surface triggers signal transduction pathways allowing the cell to sense and react to its environment in Correspondence to: Igal Szleifer. NIH Public Access Author ManuscriptLangmuir. Author manuscript; available in PMC 2010 October 20. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscriptways such as aggregating with ot...

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“…1A) to the size of the detected EVs in nm ( Fig. 1C; Fattaccioli et al, 2009;van der Pol et al, 2012).…”

Section: Introductionmentioning

confidence: 97%

Deriving Extracellular Vesicle Size From Scatter Intensities Measured by Flow Cytometry

et al. 2018

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Flow cytometry is commonly used to investigate the potential for extracellular vesicles (EVs) to be biomarkers of disease. A typical flow cytometer detects fluorescence and scatter intensities of single EVs in arbitrary units. These arbitrary units complicate data interpretation and data comparison between different flow cytometers. For example, comparison of detected EV concentrations requires knowledge of the detectable EV sizes. Using Mie theory and knowledge of the optical configuration of the flow cytometer, EV size can be derived from the scatter intensity for a given EV refractive index. Here, a protocol is described to derive the size of EVs and other nanoparticles from the scatter intensity. The resulting size distribution allows the comparison of data between flow cytometers, which is a prerequisite for clinical application of EVs as biomarkers and may advance other fields where sizing of nanoparticles is essential. © 2018 by John Wiley & Sons, Inc.

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Size and fluorescence measurements of individual droplets by flow cytometry

Cited by 42 publications

References 23 publications

Phagocytosis of immunoglobulin-coated emulsion droplets

Phagocytosis of immunoglobulin-coated emulsion droplets

Streptavidin−Biotin Binding in the Presence of a Polymer Spacer. A Theoretical Description

Deriving Extracellular Vesicle Size From Scatter Intensities Measured by Flow Cytometry

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