Umweltmedizin am IUK

Extracellular Vesicle isolation from various sources combining tangential flow filtration (TFF) and in size-exclusion chromatography (SEC), ultracentrifugation and density gradient centrifugation

We specialize in isolating extracellular vesicles from various sources, including cell cultures, organoids, bacteria, and body fluids. Our approach utilizes advanced Tangential Flow Filtration (TFF) techniques with the TFF Easy device, complemented by size-exclusion chromatography (SEC). This methodology allows us to obtain extracellular vesicles of the highest purity and integrity, suitable for subsequent analytical purposes or characterization using a range of biochemical and molecular biology techniques

Characterization of extracellular vesicles in our laboratory

Characterizing EVs can be a complex process, and we often use combination of following techniques to obtain a comprehensive understanding of their properties and functions.

Size and Concentration Analysis:

Dynamic Light Scattering (DLS): DLS measures the size distribution of EVs by analyzing how they scatter light as they move in a liquid. It provides information about the size distribution of vesicles.

Nanoparticle Tracking Analysis (NTA): NTA tracks individual EVs by analyzing the Brownian motion. It provides information about size distribution and concentration in certain size-ranges.

Morphology:

Transmission Electron Microscopy (TEM): TEM allows for direct visualization of EVs at nanoscale resolution, providing information about their shape and morphology.

Protein Composition:

Western Blotting: WB is used to detect specific proteins on the surface or within EVs, helping to identify their origin and cargo.

Mass Spectrometry: Mass spectrometry is used to analyze the protein composition of EVs in a high-throughput manner, providing detailed information about their cargo.

Protein arrays (Sciomics Gmb) are used to characterize surfaceproteins and cytokine content of EVs

MSD assay, analogous to ELISA, however several times for sensitive, is used to estimate surface EV proteins

Lipid Composition:

Lipidomics: Lipidomic analysis can identify and quantify the lipid species present in EV membranes, providing insights into their lipid composition.

Nucleic Acid Analysis:

RNA and DNA Analysis: Techniques such as RT-qPCR and Next Generation RNA sequencing  are used to analyze the RNA and DNA content of EVs, including microRNAs and other non-coding RNAs. Additionally,

Surface Marker Analysis:

Flow Cytometry: Flow cytometry is used to analyze surface markers on EVs, allowing for their phenotypic characterization.

Immunocapture Assays: Antibodies specific to EV surface markers can be used to isolate and characterize specific subpopulations of EVs.

Functional Assays:

Uptake Studies: EV uptake by target cells are assessed using techniques like confocal microscopy, which can provide insights into their functional roles in intercellular communication.