A simple immunoassay for extracellular vesicle liquid biopsy in microliters of non-processed plasma
EntityUAM. Departamento de Biología Molecular
10.1186/s12951-022-01256-5Journal of Nanobiotechnology 20.1 (2022): 72
ProjectGobierno de España. RTI2018-093569-B-I00; Gobierno de España. RED2018102411-T; Comunidad de Madrid. 2017/BMD-3733-2/IMMUNOTHERCAN
SubjectsCancer; Colloids; ELISA; Extracellular vesicles; Flocculation; Flow cytometry; Liquid biopsy; Biología y Biomedicina / Biología
Rights© The Author(s) 2022
Esta obra está bajo una licencia de Creative Commons Reconocimiento-CompartirIgual 4.0 Internacional.
Background: Extracellular vesicles (EVs), released by most cell types, provide an excellent source of biomarkers in biological fluids. However, in order to perform validation studies and screenings of patient samples, it is still necessary to develop general techniques permitting rapid handling of small amounts of biological samples from large numbers of donors. Results: Here we describe a method that, using just a few microliters of patient’s plasma, identifies tumour markers exposed on EVs. Studying physico-chemical properties of EVs in solution, we demonstrate that they behave as stable colloidal suspensions and therefore, in immunocapture assays, many of them are unable to interact with a stationary functionalised surface. Using flocculation methods, like those used to destabilize colloids, we demonstrate that cationic polymers increase EV ζ-potential, diameter, and sedimentation coefficient and thus, allow a more efficient capture on antibody-coated surfaces by both ELISA and bead-assisted flow cytometry. These findings led to optimization of a protocol in microtiter plates allowing effective immunocapture of EVs, directly in plasma without previous ultracentrifugation or other EV enrichment. The method, easily adaptable to any laboratory, has been validated using plasma from lung cancer patients in which the epithelial cell marker EpCAM has been detected on EVs. Conclusions: This optimized high throughput, easy to automate, technology allows screening of large numbers of patients to phenotype tumour markers in circulating EVs, breaking barriers for the validation of proposed EV biomarkers and the discovery of new ones
Google Scholar:Campos Silva, Carmen - Cáceres-Martell, Yaiza - Sánchez Herrero, Estela - Sandúa, Amaia - Beneitez-Martínez, Alexandra - González, Álvaro - Provencio Pulla, Mariano - Romero, Atocha - Jara-Acevedo, Ricardo - Yáñez Mo, María - Valés Gómez, Mar
This item appears in the following Collection(s)
Showing items related by title, author, creator and subject.
CtDNA from body fluids is an adequate source for EGFR biomarker testing in advanced lung adenocarcinoma
Analysis of extracellular vesicles in cancer immunomodulation and liquid biopsy Campos Silva, Carmen
mRNA in exosomas as a liquid biopsy in non-Hodgkin Lymphoma: a multicentric study by the Spanish Lymphoma Oncology Group