Circulating miRNA Extraction: A Method to Isolate miRNA from Plasma Samples Via Organic Extraction and Small RNA Enrichment

Published: April 30, 2023

Abstract

Source: Canale, M. et al., Detection of a Circulating MicroRNA Custom Panel in Patients with Metastatic Colorectal Cancer. J. Vis. Exp. (2019).

This video describes the extraction of circulating microRNA from plasma samples of colorectal cancer patients using organic extraction followed by glass-fiber column enrichment. miRNAs are stable in biological fluids such as serum and plasma, which renders them ideal circulating biomarkers for cancer diagnosis, prognosis, and treatment decision-making and monitoring. This method will help identify new circulating biomarkers capable of predicting clinical outcomes in patients.

Protocol

NOTE: Perform all of the following steps under a sterilized fume hood according to Good Laboratory Practice (GLP). 1. Plasma Collection and Storage Collect 3 mL of peripheral blood sample in a K3E EDTA tube. Centrifuge the tube at room temperature at 1,880 x g for 15 min. Recover the supernatant plasma in aliquots of 450 µL. Store the samples at -80 °C until use. NOTE:</…

Disclosures

The authors have nothing to disclose.

Materials

Rnase-free Safe-lock 1.5 mL tubes   Eppendorf 0030 123.328
Rnase-free 1000 µL tips   Starlab S1122-1830
Rnase-free Safe-lock 2 mL tubes   Eppendorf 0030 123.344
Rnase-free 20 µL tips  Starlab  S1123-1810
Rnase-free 200 µL tips  Starlab  S1120-8810
mirVana PARIS RNA and Native Protein Purification Kit   Thermo Fisher AM1556
100% ethanol anidrous ACS grade   Carlo Erba Reagents 414605
2-mercapto-ethanol   Sigma-Aldrich M3148
0.2-2, 1-10, 2-20, 20-200, 100-1000 µL laboratory pipettes
Fume hood
Benchtop microcentrifuge 
Vortex

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Cite This Article
Circulating miRNA Extraction: A Method to Isolate miRNA from Plasma Samples Via Organic Extraction and Small RNA Enrichment. J. Vis. Exp. (Pending Publication), e20331, doi: (2023).

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