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Title: Simple and Fast SEC-Based Protocol to Isolate Human Plasma-Derived Extracellular Vesicles for Transcriptional Research
Authors: Gaspar, Laetitia da Silva 
Santana, Magda M. 
Henriques, Carina 
Pinto, Maria M. 
Ribeiro-Rodrigues, Teresa M. 
Girão, Henrique 
Nobre, Rui Jorge 
Almeida, Luís Pereira de 
Keywords: biomarkers; clinical research; extracellular vesicles; plasma; size exclusion chromatography; therapy; transcriptional research
Issue Date: 11-Sep-2020
Publisher: Elsevier
Serial title, monograph or event: Molecular Therapy - Methods and Clinical Development
Volume: 18
Abstract: Extracellular vesicles (EVs) are membranous structures that protect RNAs from damage when circulating in complex biological fluids, such as plasma. RNAs are extremely specific to health and disease, being powerful tools for diagnosis, treatment response monitoring, and development of new therapeutic strategies for several diseases. In this context, EVs are potential sources of disease biomarkers and promising delivery vehicles. However, standardized and reproducible EV isolation protocols easy to implement in clinical practice are missing. Here, a size exclusion chromatography-based protocol for EV-isolation from human plasma was optimized. We propose a workflow to isolate EVs for transcriptional research that allows concomitant analysis of particle number and size, total protein, and quantification of a major plasma contaminant. This protocol yields 7.54 × 109 ± 1.22 × 108 particles, quantified by nanoparticle tracking analysis, with a mean size of 115.7 ± 11.12 nm and a mode size of 83.13 ± 4.72 nm, in a ratio of 1.19 × 1010 ± 7.38 × 109 particles/μg of protein, determined by Micro Bicinchoninic Acid (BCA) Protein Assay, and 3.09 ± 0.7 ng RNA, assessed by fluorescence-based RNA-quantitation, from only 900 μL of plasma. The protocol is fast and easy to implement and has potential for application in biomarkers research, therapeutic strategies development, and clinical practice.
ISSN: 2329-0501
DOI: 10.1016/j.omtm.2020.07.012
Rights: openAccess
Appears in Collections:I&D CNC - Artigos em Revistas Internacionais
I&D CIBB - Artigos em Revistas Internacionais
IIIUC - Artigos em Revistas Internacionais
I&D ICBR - Artigos em Revistas Internacionais
FFUC- Artigos em Revistas Internacionais

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