Separation and determination of microRNAs by high‐speed capillary sieving electrophoresis

Abstract

In this work, high-speed capillary sieving electrophoresis with laser-induced fluorescence detection was applied to simultaneously determine three microRNAs. A developed manual sample introduction device for the high-speed capillary electrophoresis system was applied to perform sample injection. Strategies, including field-amplified sample injection and electrokinetic injection, were studied to improve the detection sensitivity. Under the optimal conditions, the limit of detection for DNA-159 could be lowered to 5.10 × 10 mol/L. In order to achieve enough separation resolution, two DNA probes were designed to have extra sequences that acted as the drag tails. Under the optimized conditions, the three DNA probes and the complexes of microRNA-156, microRNA-159, and microRNA-166 could be completely separated within 3.2 min in background electrolyte (pH 8.7) containing 2.0% m/m polyvinyl pyrrolidone and 0.4% m/m hydroxyethyl cellulose. The limits of detection for the three microRNAs were 0.051, 0.11, and 0.25 nmol/L, respectively. Then the method was applied to analyze the microRNAs spiked in the samples extracted from banana leaves. The recoveries ranged from 114.3 to 121.1% (n = 3). The results showed that the method developed in this work was an effective means for microRNA assay.

Nucleic acid strand displacement for indirect determination of foodborne bacteria by capillary electrophoresis and its application in antagonism and bacteriostasis studies

“…But in this study, the longest probe 1 migrated fastest and probe 2 migrated faster than the shortest probe 3. Similar phenomena were also observed in our previous report [21]. It could be inferred that the length had minor effect on the migration speed for relatively short ssDNAs (<50 bps) modified with FAM.…”

Indirect and sensitive determination of microRNAs by magnetic field‐assisted capillary sieving electrophoresis combined with catalytic hairpin assembly

“…Each miRNA had two linear relationships in two concentration ranges. Based on three times signal‐to‐noise ratio, the measured detection limits for miRNAs were in the range of 1.12–4.05 × 10 −15 M. Without amplification, the detection limits of miRNAs, tested by CE‐LIF, could only reach the concentrations of about 0.1‐50 pM even after preconcentration (field‐amplified sample injection or isotachophoresis ) [14, 27]. Compared with the method of label‐free CE‐CHA [23], this method also exhibited higher sensitivity in the detection of miRNAs.…”

“…Separation is the typical method of the second strategy for the detection of multiple miRNAs. By separation, simultaneous detection of multiple miRNAs can be realized [13,14].…”

Microfluidics for detection of exosomes and microRNAs in cancer: State of the art

“…The detection method of laser-induced fluorescence (LIF) is a specific type of fluorescence detection, well suited for mPADs. 361,362 It should be borne in mind that the number of different intracellular miRNAs might be 1,000 or even higher. 363 Amplification or enrichment techniques play a major role in allowing miRNA detection by standard methods.…”