Extracellular vesicle (EV) analysis

One-stop-shop characterization of extracellular vesicles (EVs)

Due to their capacity to transport proteins, lipids, as well as RNA/DNA, EVs play an important role in cellular communication and are used as drug and biomarker candidates in regenerative and transplant medicine, anti-tumor therapy, and immunotherapy. TAmiRNA provides one-stop-shop services for the purification and characterization of EVs. We have established a large number of assays to study EV concentrations, size, decoration with nucleic acids and proteins, as well as DNA/RNA cargo. Our unique small RNA sequencing protocol allows determining absolute numbers of RNA molecules inside EVs. All our services are based on best practices released by the International Society for Extracellular Vesicles (ISEV).

Extracellular vesicle (EV) analysis
Extracellular vesicle (EV) analysis

benefits

EV purification: enrichment of EVs/exosomes can be performed using size exclusion chromatography (SEC), ultracentrifugation, and precipitation. SEC-based purification can deliver both EVs and protein complexes to study presence of RNA in both compartments in parallel.

EV characterization: we use both Nanoparticle Tracking Analysis (NTA Quatt) as well as multiplex flow cytometry to determine EV size, concentration, nucleic acid content (SYBR Gold), and surface epitopes (CD81, CD63, CD9). Upon request, we can develop assays for the analysis of other surface epitopes.

Non-specific characterization of EV cargo: we apply highly sensitive methods for protein, dsDNA, and ssRNA quantification in EV preparations to characterize their protein and nucleic acid content. RNase/DNase/Protease treatments are used to reduce noise from surface decorations with RNA/DNA.

exRNA analysis in EVs: we have developed robust protocols for small RNAseq, RNAseq, and RT-qPCR analysis of EVs (and protein complexes) to perform highly quantitative analysis of microRNA, mRNA, and other non-coding RNAs.

Our experience: we have completed more small RNA sequencing analyses of > 500 EV samples obtained from various biological samples. We have developed surface epitope staining protocols and have established fluorescence-based techniques for staining intracellular RNA/DNA content using NTA Quatt analysis as a means of EV manufacturing quality control.

service requirements

We can purify EVs (and protein complexes) from

  • biofluids such as (platelet-poor) plasma and urine
  • cell culture supernatant.

Sample volume is dependent on the type of biofluid and the concentration of EVs. Typically input volumes can be as low as 100 µl for plasma samples.

Alternatively, we can process EV samples that have been generated by our customers as long as certain quality parameters can be assured. Get in touch with us to discuss your specific project details.

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Comprehensive EVs characterization

Comprehensive EVs characterization

selected reference projects

  1. small RNA sequencing-based characterization of microRNAs and other ncRNAs in EVs purified from synovial fluid and plasma
  2. SEC-purification of EVs and protein complexes from 28 human urine samples (following 10 kDa-UF) and small RNA sequencing analysis.
  3. small RNA sequencing-based characterization of MSC-derived EVs intended for pharmaceutical applications.
  4. Size, concentration, and nucleic acid cargo characterization of EVs obtained from various in-vitro models and using different isolation methods.
  5. Development of a flow-cytometry based protocol to characterize oncogenic protein surface markers on EVs

publications

SVF-derived extracellular vesicles carry characteristic miRNAs in lipedema. 

Priglinger E., Strohmeier K., Weigl M et al. Sci Rep. 2020 Apr 29;10(1):7211.doi: 10.1038/s41598-020-64215-w.

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