miRNA Extraction: A Method to Extract miRNA from Plasma Sample
Published: April 30, 2023
Abstract
Source: Mensah, M. et. al., MicroRNA Based Liquid Biopsy: The Experience of the Plasma miRNA Signature Classifier (MSC) for Lung Cancer Screening. J. Vis. Exp. (2017).
MicroRNAs (miRNAs) are tissue specific, small, non-coding RNAs regulating gene expression, and could represent ideal candidates for early detection of lung cancer. In the featured protocol, we will isolate microRNA from patient blood samples for early lung cancer screening.
Protocol
1. Plasma Sample Collection Collect 10 mL of whole blood sample in Vacutainer tubes with spray-coated K2EDTA and store at room temperature. NOTE: To minimize the hemolysis, separate plasma within 2 h. Do not store the whole blood at low temperature (i.e., 4 °C) to avoid thermal shock and cell lysis that lead to an unspecific miRNA release. Within 1 h, separate the plasma by a first centrifugation step at 1,258 x g and 4 °C for 10 min. T…
Divulgaciones
The authors have nothing to disclose.
Materials
| 1x PBS | Lonza | BE-17-516 | |
| 20xTE Buffer, pH 8.0 | Molecular probe | T11493 | Dilute the 20X stock solution to have the final concentration of 0,1X in Nuclease-free water |
| Nuclease-free water | |||
| Maxwell RSC miRNA Tissue Kit | Promega | AS1460 | The kit contains Homogenization Solution, DNase I, Proteinase K, Lysis Buffer and cartridges. For 1-Thioglycerol, inhalation and contact with skin and eyes should be avoided. Use it under a Fume Cabinet. |
| Vacutainer Safety-Lock Blood Collection Set | Becton Dickinson | 367286-364815 | |
| Vortex Mixer | Velp Scientifica | F202A0173 | |
| ThermoMixer T- shaker | Euroclone | A00564 | Bring the Thermo Mixer to 37 °C before starting RNA extraction |
| Maxwell RSC Instrument | Promega | AS4500 | |
| Cryogenic vials 1.5 ml | Sigma-Aldrich | Z359033 |
Citar este artículo
miRNA Extraction: A Method to Extract miRNA from Plasma Sample. J. Vis. Exp. (Pending Publication), e20246, doi: (2023).