Extracellular vesicle microRNA quantification from plasma using an integrated microfluidic device

Abstract

Extracellular vesicles (EV) containing microRNAs (miRNAs) have tremendous potential as biomarkers for the early detection of disease. Here, we present a simple and rapid PCR-free integrated microfluidics platform capable of absolute quantification (<10% uncertainty) of both free-floating miRNAs and EV-miRNAs in plasma with 1 pM detection sensitivity. The assay time is only 30 minutes as opposed to 13 h and requires only ~20 μL of sample as oppose to 1 mL for conventional RT-qPCR techniques. The platform integrates a surface acoustic wave (SAW) EV lysing microfluidic chip with a concentration and sensing microfluidic chip incorporating an electrokinetic membrane sensor that is based on non-equilibrium ionic currents. Unlike conventional RT-qPCR methods, this technology does not require EV extraction, RNA purification, reverse transcription, or amplification. This platform can be easily extended for other RNA and DNA targets of interest, thus providing a viable screening tool for early disease diagnosis, prognosis, and monitoring of therapeutic response.

Mechanisms of Cardiovascular Disorders in Patients With Chronic Kidney Disease: A Process Related to Accelerated Senescence

“…MicroRNAs (miRNAs) are small, approximately 22-nucleotide long strands of small non-coding RNAs that regulate posttranscriptional processes (Ramshani et al, 2019;Rios et al, 2019). Unlike RNA, miRNAs show high stability in fluids and can be reliably detected in the blood (Mitchell et al, 2008).…”

“…MiRNAs levels differ in healthy and diseased tissues and play a critical role in the maintenance of cardiovascular homeostasis (Pfeifer et al, 2015). EV-associated miRNAs have been studied extensively Ramshani et al, 2019). Numerous studies have shown that miRNAs can be used as valuable pathological and therapeutic biomarkers in both cells and EVs (Bekris and Leverenz, 2015;Lei et al, 2017).…”

Sensor-Integrated Microfluidic Approaches for Liquid Biopsies Applications in Early Detection of Cancer

“…An innovative chemical-free exosome lysing system, coupled with membrane sensing, was integrated and presented by Ramshani et al [125]. The purpose of this study was to develop a PCR-free diagnostic platform in order to quantify, in a one-step manner, exosome-derived miRNA from untreated plasma samples ( Figure 5D).…”

“…The chip's detection principle relies on ion-exchange membranes, where the anionic one is functionalized with a complementary probe. The current-voltage interaction throughout the membrane sensor is drastically shifted during hybridization, which can be directly linked to miRNA levels in the sample [125].…”

Liquid biopsy technologies based on membrane microfluidics: High‐yield purification and selective quantification of biomarkers in nanocarriers

“…Such a high analyte concentration factor enhances the sensitivity of the sensors by orders of magnitude. For example, we can extend the limit of detection of our membrane sensor for microRNA from nM to pM [35,36] and for our nanoparticle sensors from 40 nM to 500 pM [37]. We shall, however, focus on how they can improve the pretreatment portion of the liquid biopsy platform here.…”