Detecting miRNA biomarkers from extracellular vesicles for cardiovascular disease with a microfluidic system

Abstract

According to World Health Organization reports, cardiovascular diseases (CVDs) are amongst the major causes of death globally and are responsible for over 18 million deaths every year. Traditional detection methods for CVDs include cardiac computerized tomography scans, electrocardiography, and myocardial perfusion imaging scans. Although diagnosis of CVDs through such bio-imaging techniques is common, these methods are relatively costly and cannot detect CVDs in their earliest stages. In contrast, the levels of certain micro RNA (miRNA) biomarkers extracted from extracellular vesicles (EVs) in the bloodstream have been recognized as promising indicators for early CVD detection. However, detection and quantification of miRNA using existing methods are relatively labor-intensive and time-consuming. In this study, a new integrated microfluidic system equipped with highly sensitive field-effect transistors (FETs) was capable of performing EV extraction, EV lysis, target miRNA isolation and miRNA detection within 5 h. The limit of detection was within the physiological range (femtomolar) for two targeted miRNAs, miR-21 and miR-126, meaning that this integrated microfluidic system has the potential to be used as a tool for early detection of CVDs.

Two-step magnetic bead-based (2MBB) techniques for immunocapture of extracellular vesicles and quantification of microRNAs for cardiovascular diseases: A pilot study

“…In this study, 2MBB was conducted with minimal special equipment required, and an overnight mixing of magnetic beads with the sample is required to attain the desired EV yield. It can be readily integrated with microfluidic systems to provide the precise and automatic control of the process and to shorten the mixing time [21,42,43]. The throughput can be further improved by utilizing field-effect transistor (FET) sensors to detect miRNAs directly without PCR amplification [44].…”

“…In order to increase the yield and purity of enriched EVs, we built on prior reported immune-affinity based techniques [14,20,21] and developed a two-step magnetic bead-based (2MBB) approach to recover EVs and EV-associated miRNAs. Micro-magnetic beads coated with capture molecules against EV-specific surface markers, such as CD63, could improve the enrichment process due to the high surface-to-volume ratio of beads.…”

On-chip miRNA extraction platforms: recent technological advances and implications for next generation point-of-care nucleic acid tests

“…Zhang et al demonstrated the usefulness of microencapsulation in combination with a particle counter for digital analysis to detect miRNAs directly from plasma in less than 3 hours and down to 10 single copies of miRNA. The device was able to detect miRNA with a limit of detection of about 50 copies per mL of plasma and was thus able to distinguish healthy donor samples from those originating from patients with metastatic colon cancer based on detection of Let-7a levels 60 magnetically attracted to the extraction solvent (blue) held by a magnetic rod (grey) 59 , c) Magnetic beads coated in anti-CD-63 antibodies for exosome isolation 61,62 , d) Surface acoustic waves for extracellular vesicle lysis 63 .…”

A Holistic Review of the State‐of‐the‐Art Microfluidics for Exosome Separation: An Overview of the Current Status, Existing Obstacles, and Future Outlook

“…Affinity‐based isolation of exosomes on chips usually relies on capture probes (typically antibodies or aptamers) to identify generic membrane markers (e.g., CD9 and CD63) or specific exosome biomarkers, such as epithelial cell adhesion molecule (EpCAM), EGFR, melanoma cell adhesion molecule, and melanoma‐associated chondroitin sulfate proteoglycan. [ 101–106 ] Compared to size‐based methods, which can only retrieve total exosomes from bio‐fluids, immunocapture on chip provides an opportunity for separating either total exosomes or sub‐type exosomes. Generally, there are two main types of immunocapture‐on‐a‐chip strategies.…”