• Implemented a novel method for transgenesis of zebrafish embryos combining microinjection and electroporation and developed a high-throughput electroporation system. • Electroporation for multiple zebrafish embryos without removing the chorion is possible by injecting genetic material inside the chorion before application of pulses. • Optimized electrical parameters (Electric pulse amplitude, number of pulses, pulse duration and pulse gap). • Optimized volume and concentration of genetic material injected inside the chorion. • The system can be converted to a high throughput automated transgenesis system for zebrafish embryos.

A schematic of the resistive-circuit method with three inlets for DI water, reagent and urine sample. The mixed fluids are transferred to a detection chamber.

Zebrafish have emerged as a useful model for biomedical research and have been used in environmental toxicology studies. However, the presence of the chorion during the embryo stage limits cellular exposure to toxic elements and creates the possibility of a false-negative or reduced sensitivity in fish embryo toxicity testing (FET). This paper presents the use of electroporation as a technique to improve the delivery of toxic elements inside the chorion, increasing the exposure level of the toxins at an early embryo stage (<3 h post-fertilization). A custom-made electroporation device with the required electrical circuitry has been developed to position embryos between electrodes that provide electrical pulses to expedite the entry of molecules inside the chorion. The optimized parameters facilitate material entering into the chorion without affecting the survival rate of the embryos. The effectiveness of the electroporation system is demonstrated using Trypan blue dye and gold nanoparticles (AuNPs, 20–40 nm). Our results demonstrate the feasibility of controlling the concentration of dye and nanoparticles delivered inside the chorion by optimizing the electrical parameters, including pulse width, pulse number, and amplitude. Next, we tested silver nanoparticles (AgNPs, 10 nm), a commonly used toxin that can lower mortality, affect heart rate, and cause phenotypic defects. We found that electroporation of AgNPs reduces the exposure time required for toxicity testing from 4 days to hours. Electroporation for FET can provide rapid entry of potential toxins into zebrafish embryos, reducing the time required for toxicity testing and drug delivery experiments.

Collecting cells from zebrafish embryos for genotyping is critical to rapid research with these model organisms. The standard collection process is manual, labor-intensive, time-consuming, and requires a skilled person to perform it. To overcome this challenge, researchers are exploring the development of automated genotyping tools for live animals, which would significantly enhance the efficiency and accuracy of genetic screening in zebrafish and other species. The focus of this research was to optimize the Zebrafish Embryo Genotyper (ZEG), an automated system used for the rapid extraction of cellular material from zebrafish embryos. This system rapidly vibrates a roughened chip containing a zebrafish embryo to collect genetic material safely and efficiently. The aim was to improve the efficiency of DNA collection from the chips used with the ZEG by identifying the key factors that contribute to the process. First, the chips were modified to resolve issues associated with loss of sample volume from the chip wells due to evaporation during processing. Second, we experimented with three critical parameters – sample volume in the wells, the vibrational frequency of the system, and the operation time – on the quantity of DNA collected. The performance was evaluated by measuring embryo survival and quantifying the DNA collected. The sensitivity (previously 90%) of the DNA collection and embryo survival (previously 95%) of the were both found to be greater than 95% after optimization. The optimized design parameters (15 µL solution volume, 2.4 V, and a 5-minute run with 5 s alternating on/off) provided a >50% increase in DNA collection compared to the previous designs and parameters. The proposed chip design and operation do not appear to cause any apparent adverse effects on the development or survival of the embryos.

This paper discusses a MEMS switch that can be fabricated using low temperature (<100^oC) deposition and patterning techniques suitable for the back-end-of-the-line integration with CMOS. The resulting cross-bar switches can be used for electrostatic discharge protection, FPGA implementation, chip security assessment and lock-down, and circuit block power management. We discuss platinum and iron switch with turn-on voltages of ~ 1.8 V. In the case of the iron switches, we also show that they can be magnetized to have “memory” and stay on when turned on. Platinum switch cycling of up to 1000 times did not show any changes in their turn-on voltage and their contact resistance was unchanged. The 10-100 nm switch airgaps were formed using low temperature sputtered sacrificial polysilicon and XeF₂ etching. XeF₂ does not attack any of the metals used in CMOS enabling fabrication of cross-bar switches with any of these metals. Once activated, it takes ∿ 6 to mechanically turn on the switch that can be decreased to ~1 ns by optimizing the device structure. Interestingly, the nm-scale gaps can be used as spark gap as a fast plasma switch to discharge first followed by the activation of the MEMS switch.