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cell electrofusion

Masa Kanduser, Mojca Kokalj Imsirovic, Marko Usaj
Induced cell fusion is a powerful method for production of hybridoma in biotechnology and cell vaccines in medical applications. Among different alternatives, physical methods have an advantage, as they do not require any additives. Among them electrofusion, an electroporation-based cell fusion method holds a great promise. Electric pulses cause cell membrane permeabilization and due to pore formation bring cell membrane into the fusogenic state. At the same time, however, they compromise cell viability. We used a train of 8 × 100 µs electric pulses, delivered at 1 Hz with strengths ranging from 400 to 1600 V/cm...
January 22, 2019: Journal of Membrane Biology
Tety Maryenti, Norio Kato, Masako Ichikawa, Takashi Okamoto
In vitro fertilization (IVF) systems using isolated gametes have been utilized to dissect post-fertilization events in angiosperms, since the female gametophytes of plants are deeply embedded within ovaries. In addition, IVF systems have been used to produce hybrid and polyploid zygotes. Complete IVF systems have been established in maize and rice, two of three major crop species providing the majority of human energy intake. Among those crop species, gametes of wheat have not been used to establish a complete IVF system successfully...
January 3, 2019: Plant & Cell Physiology
Rashmi, S Sah, S Shyam, M K Singh, P Palta
Application of cloning technology on a wide scale is severely limited by the very low live birth rate obtained with cloned embryos. Embryo quality is an important factor which affects the conception and live birth rate of cloned embryos. microRNA-21 (miR-21) has been implicated in the regulation of apoptosis and the expression level of several important genes which control apoptosis. We examined the effects of treatment of reconstructed buffalo embryos, produced by Hand-made cloning, with miR-21 mimic on developmental competence, quality and gene expression of cloned embryos...
November 26, 2018: Theriogenology
Shohei Miyakawa, Kaoru Uesugi, Keisuke Morishima
In single cell analysis, transport of foreign substances into a cell is an important technique. In particular, for accurate analysis, a method to transport a small amount (pico-liter order) of substance into the cell without leakage while retaining the cell shape is essential. Because the fusion of the cell and the giant liposome is a closed system to the outside, it may be possible to transport a precise, small amount of substances into the cell. Additionally, there is no possibility that a leaked substance would affect other systems...
July 2, 2018: Micromachines
Qingran Kong, Zhonghua Liu
The electrofusion of 2-cell embryos proves to be a simple and efficient way of generating mammalian tetraploid embryos. Many factors affect the fusion efficiency, such as fusion medium, electric field intensity, and fusion pulse length. In mice, production of tetraploid embryos by electrofusion has already been investigated; however, the investigation to produce porcine tetraploid embryos is seldom reported. In this chapter, we will describe oocytes in vitro maturation, in vitro fertilization, and the optimum conditions for electrofusion of 2-cell embryos to produce tetraploid embryos in pig...
2019: Methods in Molecular Biology
Chengxiang Li, Qiang Ke, Cheng Yao, Chenguo Yao, Yan Mi, Meng Wu, Liangpeng Ge
OBJECTIVE: Unipolar pulses have been used in cell electrofusion over the last decades. However, the problem of high mortality with unipolar pulses has not been solved effectively. The cell fusion rate is restrict by cell mortality. By using the advantages of bipolar pulses which cause less cell damage, this paper attempts to use bipolar pulses to increase the cell fusion rate. METHODS: The transmembrane voltage and pore density of cells subjected to unipolar/bipolar pulses were simulated in COMSOL software...
October 1, 2018: IEEE Transactions on Bio-medical Engineering
Monika Saini, Naresh L Selokar, Prabhat Palta, Manmohan S Chauhan, Radhey S Manik, Suresh K Singla
The first birth of a cloned animal produced through the Handmade cloning (HMC) technique was reported more than 15 years ago in cattle. This method of somatic cell nuclear transfer (SCNT) has subsequently been evolving as a much simpler alternative to the classical micromanipulator-based SCNT. Several farm animal species such as cattle, buffalo, pigs, sheep, and goats have been successfully cloned using HMC. In buffalo, HMC technique is now well established, and several births of cloned calves have been reported by us...
August 9, 2018: Animal Reproduction Science
A Davari, M Skinner, B L Parker
AIMS: The aim of this study was to optimize the potential of Beauveria bassiana through cell electrofusion technique with emphasis on improving its thermotolerance and virulence against western flower thrips, Frankliniella occidentalis (Thysanoptera: Thripidae) (WFT). METHODS AND RESULTS: Nine B. bassiana isolates were tested against western flower thrips, F. occidentalis (Thysanoptera: Thripidae) (WFT) to estimate their pathogenicity and speed of infection under laboratory conditions...
June 25, 2018: Journal of Applied Microbiology
Chengxiang Li, Qiang Ke, Chenguo Yao, Yan Mi, Hongmei Liu, Yanpeng Lv, Cheng Yao
Traditionally, microsecond pulsed electric field was widely used in cell electrofusion technology. However, it was difficult to fuse the cells with different sizes. Because the effect of electroporation based on microsecond pulses was greatly influenced by cell sizes. It had been reported that the differences between cell sizes can be ignored when cells were exposed to nanosecond pulses. However, pores induced by those short nanosecond pulses tended to be very small (0.9 nm) and the pores were more easy to recover...
2018: PloS One
Kumari Priti Sinha, Rochish M Thaokar
Vesicles or biological cells under simultaneous shear and electric field can be encountered in dielectrophoretic devices or designs used for continuous flow electrofusion or electroporation. In this work, the dynamics of a vesicle subjected to simultaneous shear and uniform alternating current (ac) electric field is investigated in the small deformation limit. The coupled equations for vesicle orientation and shape evolution are derived theoretically, and the resulting nonlinear equations are handled numerically to generate relevant phase diagrams that demonstrate the effect of electrical parameters on the different dynamical regimes such as tank treading (TT), vacillating breathing (VB) [called trembling (TR) in this work], and tumbling (TU)...
March 2018: Physical Review. E
Rogier M Schoeman, Wesley T E van den Beld, Evelien W M Kemna, Floor Wolbers, Jan C T Eijkel, Albert van den Berg
We present a microfluidic chip that enables electrofusion of cells in microdroplets, with exchange of nuclear components. It is shown, to our knowledge for the first time, electrofusion of two HL60 cells, inside a microdroplet. This is the crucial intermediate step for controlled hybridoma formation where a B cell is electrofused with a myeloma cell. We use a microfluidic device consisting of a microchannel structure in PDMS bonded to a glass substrate through which droplets with two differently stained HL60 cells are transported...
February 27, 2018: Scientific Reports
Girja S Shukla, Stephanie C Pero, Yu-Jing Sun, Chelsea L Carman, Seth Harlow, David N Krag
Autoantibodies to breast and other cancers are commonly present in cancer patients. A method to rapidly produce these anti-cancer autoantibodies in the lab would be valuable for understanding immune events and to generate candidate reagents for therapy and diagnostics. The purpose of this report is to evaluate sentinel nodes (SNs) of breast cancer patients as a source of anti-cancer antibodies. Radiotracer lymphatic mapping in 29 patients with breast cancer confirmed the identity of the SNs which provided source cells for this study...
April 2018: Journal of Immunological Methods
Tina Batista Napotnik, Gianluca Bello, Eva-Kathrin Sinner, Damijan Miklavčič
Polymersomes, vesicles composed of block copolymers, are promising candidates as membrane alternatives and functional containers, e.g., as potential carriers for functional molecules because of their stability and tunable membrane properties. In the scope of possible use for membrane protein delivery to cells by electrofusion, we investigated the cytotoxicity of such polymersomes as well as the effects of nanosecond electric pulses with variable repetition rate on the shape and permeability of polymersomes in buffers with different conductivities...
October 2017: Journal of Membrane Biology
R Eid, U Petzoldt
Asynchronous tetraploid mouse embryos were generated by electrofusion of fertilized eggs with blastomeres from different cleavage stages. The majority of the cytoplasm was always contributed by the egg. The best development was observed when eggs were fused with 2-cell blastomeres. Both genomes became active in fusion embryos (at least the genes for glucose phosphate isomerase did). Stage-specific protein synthesis seemed to be more adjusted to the developmental stage of the egg's than of the blastomere's genome, but at the 2-cell stage both contributed slightly differently to the protein patterns...
April 1993: Roux's Archives of Developmental Biology: the Official Organ of the EDBO
Lei Cheng, Haizhen Wu, Jing Fei, Lujia Zhang, Jiang Ye, Huizhan Zhang
Objective To prepare and characterize a monoclonal antibody (mAb) against 4-amylphenol (4-AP), clone its cDNA sequence and make homology modeling for its Fv fragment. Methods A high-affinity anti-4-AP mAb was generated from a hybridoma cell line F10 using electrofusion between splenocytes from APA-BSA-immunized mouse and Sp2/0 myeloma cells. Then we extracted the mRNA of F10 cells and cloned the cDNA of mAb. The homology modeling and molecular docking of its Fv fragment was conducted with biological software...
March 2017: Xi Bao Yu Fen Zi Mian Yi Xue za Zhi, Chinese Journal of Cellular and Molecular Immunology
A Denzi, J A A Escobar, C Nasta, C Merla, B Benassi, C Consales, F Apollonio, M Liberti
In the last decades, the effects of ultrashort pulsed electric fields have been investigated demonstrating their capability to be involved in a great number of medical applications (e.g. cancer, gene electrotransfer, drug delivery, electrofusion). In particular, experiments in literature demonstrate that internal structures can be involved when pulse duration is reduced. Up to now, the mechanism that permits the electroporation phenomenon has not been completely understood and hence atomistic, microdosimetry and dosimetry models have been developed to help in this field...
August 2016: Conference Proceedings: Annual International Conference of the IEEE Engineering in Medicine and Biology Society
Yong Guo, Hejuan Li, Ying Wang, Xingrong Yan, Xihui Sheng, Di Chang, Xiaolong Qi, Xiangguo Wang, Yunhai Liu, Junya Li, Hemin Ni
Somatic cell nuclear transfer (SCNT) is frequently used to produce transgenic cloned livestock, but it is still associated with low success rates. To our knowledge, we are the first to report successful production of transgenic cattle that overexpress bovine adipocyte-type fatty acid binding proteins (A-FABPs) with the aid of SCNT. Intragenomic integration of additional A-FABP gene copies has been found to be positively correlated with the intramuscular fat content in different farm livestock species. First, we optimized the cloning parameters to produce bovine embryos integrated with A-FABP by SCNT, such as applied voltage field strength and pulse duration for electrofusion, morphology and size of donor cells, and number of donor cells passages...
February 2017: Molecular Biology Reports
S Sakamoto, K O Okeyo, S Yamazaki, O Kurosawa, H Oana, H Kotera, M Washizu
Although fusion of somatic cells with embryonic stem (ES) cells has been shown to induce reprogramming, single-cell level details of the transitory phenotypic changes that occur during fusion-based reprogramming are still lacking. Our group previously reported on the technique of one-to-one electrofusion via micro-slits in a microfluidic platform. In this study, we focused on developing a novel air-lock patterning technique for creating localized adhesion zones around the micro-slits for cell localization and real-time imaging of post fusion events with a single-cell resolution...
September 2016: Biomicrofluidics
Jeong Ho Hwang, Sang Eun Kim, Mukesh Kumar Gupta, HoonTaek Lee
Transgenic animal producing technology has improved consistently over the last couple of decades. Among the available methods, somatic cell nuclear transfer (SCNT) technology was officially the most popular. However, SCNT has low efficiency and requires a highly skilled individual. Additionally, the allo-SCNT nuclear reprogramming mechanism is poorly understood in the gnotobiotic miniature pig, which is a candidate for xenotransplantation, making sampling in oocytes very difficult compared to commercial hybrid pigs...
August 2016: Cellular Reprogramming
Linying Liu, Zheng Mao, Jianhua Zhang, Na Liu, Qing Huo Liu
The effects of electric field on lipid membrane and cells have been extensively studied in the last decades. The phenomena of electroporation and electrofusion are of particular interest due to their wide use in cell biology and biotechnology. However, numerical studies on the electrofusion of cells (or vesicles) with different deformed shapes are still rare. Vesicle, being of cell size, can be treated as a simple model of cell to investigate the behaviors of cell in electric field. Based on the finite element method, we investigate the effect of vesicle shape on electrofusion of contact vesicles in various medium conditions...
2016: PloS One
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