GSE122068
Comparison of methods for the characterization of miRNAs from 200 µl of cerebrospinal fluid
Cerebrospinal fluid (CSF) miRNAs have emerged as a potential low invasive diagnostic tool for central nervous system malignancies. However, they have not yet been implemented in the clinic since there is no standardized and simple method established. Another problem that it must be considered is that the amount of CSF that can be obtained from patients is very limited, especially from infants. With this in mind, we have compared six different methodologies to establish the best one for the detection of miRNAs using a minimum CSF volume. Four of them incorporated a EVs enrichment step and the other two extracted the miRNAs directly from crude CSF. The efficiency of each method was evaluated by quantitative PCR (qPCR) and smallRNA sequencing. Our results showed that, the Plasma/Serum RNA Purification kit (Norgen) and the proposed method combining the total exosome isolation reagent (Invitrogen) with the mirVana PARIS kit (Ambion) were the most efficient. However, the Norgen's kit was the most reproducible protocol by qPCR and presented a user-friendly protocol. Therefore, we considered it as the most suitable method for detecting circulating and EVs-associated miRNAs using only 200 µl of CSF. Overall design: Comparison of six methodologies for miRNAs analysis from 200 µl of cerebrospinal fluid. Of them, four incorporated a EVs enrichment step and the other two extracted the miRNAs directly from crude CSF. The efficiency of each method was evaluated by quantitative PCR (qPCR) and smallRNA sequencing. For this experiment a pool of samples was generated.