New and improved methods for characterization of tumour-specific markers in exosomes

Background: The activation of the immune system mediated by engagement of NKG2D with its ligands is a crucial step in the regulation of both innate and specific immune responses, in particular, in immune recognition of cancer. NKG2D-ligand expression is upregulated when the cells suffer different ty...

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Bibliographic Details
Published in:Journal of extracellular vesicles 2018-01, Vol.7, p.127-127
Main Authors: Silva, Carmen Campos, Cobo, Sheila López, Montero, Henar Suárez, Artime, Amanda Moyano, Paschen, Annette, Blanco-López, Maria del Carmen, Yáñez-Mó, María, Valés, Mar
Format: Article
Language:English
Subjects:
Antibodies
Biomarkers
Cancer
Enzyme-linked immunosorbent assay
Exosomes
Flow cytometry
Immune response
Ligands
Tumors
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Summary:Background: The activation of the immune system mediated by engagement of NKG2D with its ligands is a crucial step in the regulation of both innate and specific immune responses, in particular, in immune recognition of cancer. NKG2D-ligand expression is upregulated when the cells suffer different types of stress, notably tumoural transformation. However, the NKG2D-mediated response can be modulated by the release of these molecules to the extracellular milieu, leading to immune evasion. Methods: To generate tools that facilitate investigation of the role of the presence of the NKG2D-ligand MICA in tumour derived-exosomes, we have analysed different methods for the detection and characterization of tumour-derived exosomes including newly developed lateral flow devices and bead capture-based flow cytometry tests. Results: Comparison of the different techniques; Western blot, ELISA, flow cytometry and lateral flow, demonstrates that the use of the same combination of tetraspanins and tumour markers antibodies can result in very different outcomes when using different techniques. In fact, when optimising the combinations and concentrations of antibodies for use in each technique, special care had to be taken due to the risk of multimeric aggregate formation. Summary/Conclusion: Translating methods originally established for detection of soluble molecules into the detection of vesicles needs a careful optimization of each technique. The implications of these data for the detection of tumour markers in exosomes of biological samples will be discussed.
ISSN:2001-3078