Abstract
The preservation of nucleic acids from clinical samples is critical to facilitate accurate molecular diagnosis. The use of a paper matrix, Flinders Technology Associates (FTA) Elute cards, to archive DNA and viral RNA is well-documented. However, the feasibility of FTA Elute cards for archiving serum and serum exosomal microRNAs (miRNAs) remains unclear. Here, we performed a comprehensive evaluation of FTA Elute cards for miRNA storage and recovery in different pre-analytical conditions. The recovery of serum miRNA dry-spotted on FTA Elute cards by direct elution with water at high temperature was poor. However, serum miRNAs dry-spotted on the cards were isolated with about 40% yield when using QIAzol lysis reagent and recovery was improved remarkably (>80%) upon extraction from cards pre-treated with trehalose. miRNAs stored on the cards remained stable at room temperature and can be kept for prolonged periods. Furthermore, miRNAs could be similarly recovered from serum exosomes dry-spotted on the cards. Importantly, when using sera from gastric cancer (GC) patients, the miRNAs were efficiently recovered from trehalose pre-treated cards without affecting their representation. Collectively, we have demonstrated the potential of FTA Elute cards to archive serum and serum exosomal miRNAs, making it useful for biomarker discovery and diagnostics.
Highlights
MicroRNAs are small non-coding RNAs (19–22 nucleotides) that regulate protein expression and are physiologically significant in several cellular processes[1,2]
We investigated the ability of Flinders Technology Associates (FTA) Elute cards to archive and subsequently retrieve serum exosomal miRNAs. miRNAs recovered from dry-spotted serum exosomes on FTA Elute cards using QIAzol lysis reagent were compared with miRNAs extracted directly from serum exosomes, serving as controls
FTA Elute cards have been successfully used for the collection and storage of viral RNA, and DNA from biological samples[21,22,23,24], there are instances where the recovery of nucleic acids was unacceptably low, as with enteroviral RNA (~6.1%)[24]
Summary
MicroRNAs (miRNAs) are small non-coding RNAs (19–22 nucleotides) that regulate protein expression and are physiologically significant in several cellular processes[1,2]. Unlike other forms of RNA, miRNAs are remarkably stable in clinical samples like plasma and serum In these biofluids, miRNAs are protected from degradation by a) encapsulation in microvesicles or exosomes[3], b) binding to high-density lipoproteins[4] or c) binding to the Argonaute 2 protein complex (AGO2)[5]. The Flinders Technology Associates (FTA) Elute cards (GE Healthcare) have been used successfully to collect, transport and archive nucleic acids of various pathogens, which include viral DNA and RNA21–24. We recovered serum and serum exosomal miRNAs from FTA Elute cards using a commercially available reagent, QIAzol lysis reagent This method is efficient without the need to heat the samples. We found that FTA Elute cards pre-treated with trehalose significantly enhanced the recovery of these miRNAs
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