Currently available methods for cell separation are generally based on fluorescent labeling using either endogenously expressed fluorescent markers or the binding of antibodies or antibody mimetics to surface antigenic epitopes. However, such modification of the target cells represents potential contamination by non-native proteins, which may affect further cell response and be outright undesirable in applications, such as cell expansion for diagnostic or therapeutic applications, including immunotherapy. We present a label- and antibody-free method for separating macrophages from living Drosophila based on their ability to preferentially phagocytose whole yeast glucan particles (GPs). Using a novel deswelling entrapment approach based on spray drying, we have successfully fabricated yeast glucan particles with the previously unachievable content of magnetic iron oxide nanoparticles while retaining their surface features responsible for phagocytosis. We demonstrate that magnetic yeast glucan particles enable macrophage separation at comparable yields to fluorescence-activated cell sorting without compromising their viability or affecting their normal function and gene expression. The use of magnetic yeast glucan particles is broadly applicable to situations where viable macrophages separated from living organisms are subsequently used for analyses, such as gene expression, metabolomics, proteomics, single-cell transcriptomics, or enzymatic activity analysis.

• Klíčová slova
• cell separation, iron oxide nanoparticles, phagocytosis, spray drying, β-glucan particles,
• MeSH
• Drosophila melanogaster metabolismus   MeSH
• glukany * chemie   metabolismus   MeSH
• magnetické jevy MeSH
• makrofágy metabolismus   MeSH
• Saccharomyces cerevisiae * chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glukany * MeSH

Microfluidic devices, allowing superior control over the spatial and temporal distribution of chemical substances and high process reproducibility, are nowadays essential in various research areas and industrial fields where the traditional "macroscopic" approach was no longer able to keep up with the increasing demands of high-end applications. In the present work, internal mixing of droplets formed by a flow-focusing X-junction at constant flow rates of both phases for three different channel heights (i.e. 20, 40 and 60 μm) was investigated and characterised. Both experimental methods and 3D CFD simulations were employed in order to resolve governing factors having an impact on internal mixing and homogenization time of model tracers inside of droplet reactors. Additionally, the influence of channel height on internal mixing was experimentally studied for continuous preparation of iron oxide nanoparticles by co-precipitation reaction. Since the initial nucleation phase is strongly affected by mixing and spatial distribution of all reactants, the final particle size and particle size distribution (PSD) can be used as direct indicators of mixing performance. It has been demonstrated that the smallest 20 μm channels provided narrower PSD and smaller particle mean size compared to higher channels.

• Publikační typ
• časopisecké články MeSH