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 |
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).