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Isolation and Characterization of Tissue-Derived Extracellular Vesicles from Mouse Lymph Nodes.

Bernadett R Bodnár^1,2,3, Sayam Ghosal^1,2, Brachyahu M Kestecher^1,2,3

¹Institute of Genetics, Cell- and Immunobiology, Semmelweis University, 1089 Budapest, Hungary.

International Journal of Molecular Sciences

|July 12, 2025

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Researchers developed a new method to isolate extracellular vesicles (EVs) from mouse lymph nodes. Immunization significantly altered these EVs, offering insights into immune responses.

Area of Science:

Immunology
Cell Biology
Biochemistry

Background:

Extracellular vesicles (EVs) are crucial intercellular communicators released by cells.
Isolating and characterizing EVs from specific tissues like lymph nodes (LNs) is vital for understanding immune responses.
Existing methods may not fully capture the complexity of EVs in lymphoid tissues.

Purpose of the Study:

To establish a robust protocol for isolating and characterizing EVs from mouse lymph nodes.
To investigate the impact of immunization on the properties of lymph node-derived EVs.
To analyze changes in immune and EV markers following immunization.

Main Methods:

EV isolation from mouse lymph nodes using differential centrifugation and size-exclusion chromatography.

Keywords:

exosomes extracellular vesicles immunization lymph node ovalbumin

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Morphological characterization via transmission electron microscopy.

Quantification and characterization using nanoparticle tracking analysis, bicinchoninic acid assay, sulfo-phospho-vanillin assay, flow cytometry, and Western blot.

Main Results:

Successful isolation and characterization of both small and large EVs from mouse LNs.
Demonstrated significant alterations in EV particle concentration, size distribution, protein-to-lipid ratio, and marker expression post-immunization.
Identified distinct changes in immune and EV markers in response to immunization.

Conclusions:

A novel and effective protocol for isolating and characterizing EVs from lymph nodes has been established.
Immunization significantly impacts the physical and molecular properties of lymph node EVs.
These findings provide valuable insights into the role of EVs in immune system modulation and response.