ELECTROPHORESIS最新文献

This work investigates electrokinetically driven protein transport in open microfluidic devices with microwells featuring axial shape variations. The results indicate that protein propagation, which is lysed at the surface, depends on two key parameters: the electrical potential ratio $��\left(��\alpha �=�\zeta �/�{\varphi }_0��\right)�$, and the geometric curvature of the microwell. The concave microwell configuration presents the best outcome due to the emergence of a transverse velocity component that confines the cell within the microwell. Lastly, protein concentration can be improved when the negative microwell geometric slope exhibits nondifferentiable behavior (e.g., edges or fractal geometries), while a higher zeta potential can broaden the influence of the Stern layer.

Dielectrophoresis (DEP) is a powerful tool for manipulating particles using non-uniform electric fields. This study combines numerical simulations and experiments to investigate crossover frequencies (COFs) for micro- and nanoparticles in a 3D microfluidic device with circular traps. MATLAB simulations revealed an inverse relationship between particle size and COF. For microparticles with diameters of 1.03, 2.27, 4.42, and 6.83 µm, the COFs were calculated as 769.10, 352.76, 183.96, and 120.51 kHz, respectively. For nanoparticles measuring 50, 170, and 500 nm, the corresponding COFs were 15.6, 4.62, and 1.57 MHz. These results closely matched experimental data. Notably, additional low-frequency pseudo-COFs emerged in experiments for nanoparticles ranging from 2 to 8 kHz (50 nm), 10 to 50 kHz (170 nm), and 40 to 100 kHz (500 nm). These frequencies proportionally increased with nanoparticle size and corresponded to unexpected negative DEP (nDEP)-like behavior under positive DEP (pDEP) conditions. This effect is attributed to low-frequency alternating current electroosmosis (ACEO), which dominates the DEP response of the nanoparticles smaller than 1 µm. These findings demonstrate strong agreement between numerical simulations and experimental results while also revealing the limitations of traditional models in predicting nanoparticle behavior under DEP. We expect that these results can also be applied to the manipulation of various bioparticles.

Brucellosis is a neglected zoonotic disease that continues to impact global public health and livestock economies, particularly in regions with limited diagnostic infrastructure. Its causative agent, Brucella abortus, is difficult to detect due to its intracellular lifestyle and the nonspecific symptoms it causes in humans. This study demonstrates the experimental application of dielectrophoresis (DEP) in a microfluidic device for the selective manipulation of polystyrene beads and inactivated B. abortus bacteria. By tuning the frequency and medium conductivity, reliable combined negative dielectrophoretic (nDEP) and hydrodynamic flow responses were achieved, leading to the deflection of bacterial cells across the microchannel within a critical vertical window for particle control. Distinct particle trajectories were observed under varying electric field conditions, confirming effective separation without the need for labels or biochemical markers, except for visual validation. This label-free strategy enables rapid sample processing and has the potential to be integrated into portable platforms for on-site diagnostics. The results highlight the feasibility of DEP-based approaches for pathogen separation and support their future implementation in brucellosis surveillance and point-of-care testing.

CRISPR-Cas9-targeted sequencing can enrich DNA regions of interest by directing the Cas9 protein to bind and cleave specific DNA sequences via single-guide RNA (sgRNA). It is interesting to explore the efficacy of using CRISPR-Cas9-targeted nanopore sequencing (referred to as Cas9-seq), a polymerase chain reaction (PCR)-free workflow, for forensic short tandem repeats (STR) profiling, and to compare it with the amplification-based approach. In this pilot study, we constructed a Cas9-seq method for profiling seven STR loci, including D18S51, FGA, TPOX, D16S539, vWA, CSF1PO, and TH01. With 3 µg DNA inputs from human NA12878 and 293T cell lines, we achieved 643.45- and 468.34-fold enrichment ratios of the sgRNA-targeted regions by using Cas9-seq, respectively. Compared to nanopore sequencing of PCR amplicon products (amplicon-seq) of the ForenSeq DNA Signature Prep kit, the Cas9-seq reads had an ultralow strand bias. However, surprisingly, Cas9-seq did not show advantages in allele balance and had higher noise in the reads. At the seven STR loci for the two samples, both Cas9-seq and amplicon-seq had three genotyping errors. Additionally, there were no false-positive single-nucleotide polymorphisms (SNPs) introduced by Cas9-seq, whereas amplicon-seq produced three. In sum, we conclude that the PCR-free Cas9-seq might not be favorable for forensic STR genotyping.