Tza-Huei Wang

Microspore‐derived embryogenesis (MDE) is essential for doubled haploid (DH) plant breeding, but conventional bulk culture systems are limited by poor scalability and difficulty for automation. Adapting droplet microfluidics for plant cell culture is promising but challenging due to the large size and fragility of plant cells involved in embryogenesis, such as maize microspores (60–85 µm) and their expanding embryo‐like structures (200–500 µm). Here, we present a droplet microfluidic platform engineered for large plant cells, integrating a tubing‐based loading strategy that minimizes sedimentation losses, robust generation of stable 20 and 200 nL droplets optimized for large‐cell encapsulation, and scalable 3D reservoir incubation for long‐term culture. Optimized flow conditions and surface treatments enabled cell encapsulation with high fidelity, sustaining viability for over two weeks. Multi‐modal viability assessment using fluorescence spectroscopy and microscopy (bright‐field and fluorescence) confirmed that droplet culture preserves cell health comparably to bulk methods. The platform supports both single‐cell and few‐cell encapsulation modes, and increasing droplet volume and cell loading density further enhances embryogenesis rates and total embryo yield, respectively. Together, these advances establish a scalable, modular droplet microfluidic platform for high‐throughput embryogenesis analysis and embryo production, offering new opportunities in plant developmental biology, precision breeding, and doubled‐haploid technology.

Alterations of the epigenome, including histone tail post-translational modifications (PTMs), have been shown to regulate the expression of genes important for driving multiple cellular processes including cellular differentiation, aging and tumorigenesis. Although the ability to measure histone tail PTM levels is integral to the understanding of chromatin biology, because of technological limitations, the proportion of nucleosomes that have histone tail PTMs at genomic loci remain undefined. To overcome this limitation, we developed the i mmuno-profiling of d Ca s9 t argeted ch romatin (I-CATCH) system, a technology designed to measure and normalize the levels of nucleosomes and co-occurring histone tail PTMs at targeted loci. Here, we show that the I-CATCH system has high analytical performance for measuring the levels of histone tail PTMs and we report the normalized levels of multiple histone tail PTMs at different genomic loci from both HeLa cells and human plasma. While further studies are required to determine the full utility of the I-CATCH system, we are hopeful that the I-CATCH system will help enable the ability to investigate new important features of chromatin biology.

Early detection of cancer remains essential for improving patient outcomes, especially in resource-constrained areas that lack access to standard testing. Point-of-care (POC) detection of DNA methylation biomarkers has emerged as a promising solution, but fully automated testing platforms have yet to be demonstrated. Herein we report a portable and autonomous platform for POC detection of DNA methylation biomarkers. This platform integrates droplet magnetofluidic sample processing and on-cartridge wax valves to eliminate operator intervention midway and facilitate walk-away convenience. Moreover, imaging-based optical detection, in tandem with a four-channel cartridge design, enables parallelized TaqMan probe-based methylation-specific qPCR for the simultaneous detection of three DNA methylation biomarkers and an assay control gene. Focusing on the use case of cervical cancer—the fourth most common cancer in women worldwide—this platform was equipped to detect methylation biomarkers AJAP1, EPO, and HS3ST2 from HeLa S3 cervical adenocarcinoma (CAC) cells. This platform was further tested with contrived cervical cancer samples by spiking HeLa S3 CAC cells in archived normal cervix Papanicolaou (Pap) specimens (n = 6). In conclusion, this platform demonstrates automated POC detection of cervical cancer methylation biomarkers and may emerge as an accessible tool for early cancer detection in resource-constrained settings.

Scalable, cost-effective manufacturing remains a major barrier to the clinical translation of viral vector–mediated gene therapies. The widely used transfection method for producing adeno-associated virus (AAVs) and lentivirus uses plasmid DNA (pDNA)/polyethyleneimine (PEI) particles loaded with multiple plasmids; however, these particles must be prepared at low concentrations and used immediately, limiting scalability and reproducibility. Here we show a kinetic-gating strategy in which transient binding of trivalent citrate ions slows complexation, modulating charge-neutralization kinetics and delaying particle nucleation, enabling the formation of stable, highly concentrated pDNA/PEI particles. By incorporating citrate, we prevent aggregation and achieve uniform assembly at high concentrations, enabling a ten-fold increase in DNA concentration (to 0.2 mg/mL) and reduced dosing volumes. The method is robust across mixing conditions, compatible with standard manufacturing workflows, and maintains AAV production efficiency across scales. These results establish a simple and generalizable approach to control polyelectrolyte assembly kinetics, improving the scalability and reliability of viral vector production.

Paramagnetic bead–based capture and release is widely used for nucleic acid preparation in benchtop and automated workflows. In emerging magnetofluidic and cartridge-style point-of-care systems, sequential bead elution has only recently begun to be explored as a strategy to distribute limited nucleic acids across multiple reactions. In such designs, elution behavior becomes directly relevant to assay performance and multiplexing capacity. Here, we systematically examine multi-sequential elutions from silica paramagnetic beads and quantify each eluate by qPCR. We describe the data with a phenomenological, concentration-dependent, exponential-decay model and show that elution profiles depend on nucleic acid target size (soybean genomic DNA versus a short synthetic double-stranded DNA fragment, gBlock), starting concentration, and multiplexed (duplex) bead binding. We demonstrate reproducible nucleic acid signal across eight elutions, extract size-dependent decay constants, and evaluate model performance over a range of input concentrations. Duplex and alternating-trait experiments further show that the same framework can describe multi-target elution with limited data points per target. Together, these results provide a quantitative description of sequential nucleic acid elution from silica beads and a simple modeling approach that can inform the design of multi-step bead-based workflows.

Simultaneous quantification of multiple biomarker classes from blood is essential for improving diagnostic accuracy, yet current methods remain limited by low multiplexing capacity, reliance on separate instruments and workflows, and inconsistent signal readouts that hinder data integration. Here, we present fluorescence-coded, magnetic bead-enhanced digital co-detection of multianalytes (FluoMag-dCoDe), a platform that converts diverse analytes into distinct fluorescence-coded DNA templates for multiplexed quantification in a single digital polymerase chain reaction (dPCR) using a universal primer and TaqMan probe set. We demonstrate comeasurement of eight mRNA and three protein immune-response biomarkers in a two-color dPCR format. The platform achieves high quantitative fidelity with minimal cross-interference, showing strong concordance with single-plex RT-qPCR for mRNAs and superior analytical sensitivity to ELISA for proteins from ∼35 μL fingerstick-compatible blood volumes. By unifying mRNA and protein quantification in a single standardized readout while sustaining robust performance from low blood volumes, FluoMag-dCoDe establishes a scalable approach for high-dimensional biomarker analysis in precision diagnostics.

The ability to accurately measure aberrant DNA methylation levels is integral to the understanding of DNA methylation biology. It is well-established that in cancer, the largest, and thus, most biologically important absolute gains of DNA methylation levels occur at CpG sites with low native levels while the largest losses occur at CpG sites with high native levels. Conventional wisdom assumes that the observed association between the degree of the alterations and the native levels are largely due to the limitations of change within the DNA methylation scale. Here, we present evidence that this association is largely caused by alterations occurring as a global rate of change relative to the native level. We show that DNA methylation alterations can be accurately compared by calculating the rate of change relative to the native level. Most importantly, this approach enables the identification of more biologically significant DNA methylation alterations.