600 µL of cell supernatants and rat plasma were centrifuged (10,000 × g for 30 min at 4 °C) to remove cell debris, and the supernatant was recovered. For RNA isolation of EVs the exoRNeasy serum/plasma midi kit (Qiagen; Hilden, Germany) was used. During the RNA purification step, the same amount of cel-miR-39 spike-in control was added (Qiagen; Hilden, Germany) according to the provider recommendations and previous publication (Enderle et al., 2015 (link)). The RNA isolated from EVs was immediately converted to cDNA, as described below.
Hernández-Díazcouder A., González-Ramírez J., Giacoman-Martínez A., Cardoso-Saldaña G., Martínez-Martínez E., Osorio-Alonso H., Márquez-Velasco R., Sánchez-Gloria J.L., Juárez-Vicuña Y., Gonzaga G., Sánchez-Lozada L.G., Almanza-Pérez J.C, & Sánchez-Muñoz F. (2021). High fructose exposure modifies the amount of adipocyte-secreted microRNAs into extracellular vesicles in supernatants and plasma. PeerJ, 9, e11305.
Other organizations :
Universidad Autónoma Metropolitana, Instituto Nacional de Cardiología, Universidad Autónoma de Baja California, Instituto de Medicina Genómica, National Institute of Genomic Medicine, Instituto Nacional de Cardiologia
Addition of cel-miR-39 spike-in control during RNA purification step
positive controls
Cel-miR-39 spike-in control
negative controls
Not explicitly mentioned
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