Svirshchevskaya, E. V. | bioRxiv Science

bioRxiv2021

Asymmetric depth-filtration - a versatile and scalable approach for isolation and purification of extracellular vesicles

A novel asymmetric depth filtration (DF) approach for isolation of extracellular vesicles (EVs) from biological fluids is presented, and its performance is compared with established methods. The developed workflow is simple, inexpensive, and relatively fast. Compared with ultracentrifugation and size-exclusion chromatography, the developed method isolates EVs with higher purity and yield. Only standard laboratory equipment is needed for its implementation, which makes it suitable for low-resource locations. The described implementation of the method is suitable for EV isolation from small biological samples in diagnostic and treatment guidance applications. Following the scale-up routes adopted in the biomanufacturing of therapeutics, which routinely rely on DF as one of the product purification steps, the developed method may be scaled up to harvesting therapeutic EVs from large volumes of growth medium.

bioengineering↗

bioRxiv2021

The IgE production is initially induced in subcutaneous fat and depends on extrafollicular B cells

BackgroundGrowing body of evidence indicates that IgE production can be developed by mechanisms that differ from those responsible for IgG and IgA production. One potential possibility is generation of IgE producing cells from tissue-associated B-cells and/or through extrafollicular pathway. But the role of subcutaneous fat-associated B-cells in this process is poorly investigated. The aim of the present study was to investigate the role of different B- and T- cell subpopulations after long-term antigen administration in IgE response. MethodsBALB/c mice were immunized 3 times a eeks for 4 weeks in withers region enriched with subcutaneous fat with high and low antigen doses as well as by intraperitoneal route in region enriched with visceral fat for comparison. ResultsAfter long-term antigen administration that promotes the type of immune response which is more similar to one observed in young allergic children, subcutaneous fat tissue B-cells generates more rapid and active IgE class switched and IgE-produced cells. Although IgE production at later time points was initiated also in regional lymph nodes, the early IgE production was exclusively linked with subcutaneous fat. We observed that low-dose induced strong IgE production accompanied by minimal IgG1 production was linked with extrafollicular B-2 derived plasmablasts as well as extrafollicular T- helpers accumulation. Delayed IgE class switching in regional lymph nodes and visceral fat tissue was characterized by the absence of both stable plasmablasts and T-extrafollicular helpers accumulation. ConclusionExtrafollicular B- and T-cell responses in subcutaneous fat are necessary for early IgE class switching and sensitization process in the case of allergen penetration through skin.

immunology↗