Academic literature on the topic 'Protein electroporation'
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Journal articles on the topic "Protein electroporation"
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Berneman, Zwi N., Evelien Smits, Peter Ponsaerts, Marc Lenjou, Griet Nijs, Dirk R. Van Bockstaele, and Viggo F. Van Tendeloo. "RNA Electroporation as a New Gene Transfer Method in Hematopoietic Progenitor Cells, Mesenchymal Cells and Activated T-Cells." Blood 104, no. 11 (November 16, 2004): 5269. http://dx.doi.org/10.1182/blood.v104.11.5269.5269.
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Abstract Electroporation of messenger RNA has become an established method for gene transfer into dendritic cells for immunotherapeutic purposes. When electroporating dendritic cells, this method combines high transient gene expression with low cell mortality without the risk for insertional mutagenesis. Here, we have investigated the potential of mRNA electroporation to induce short-term transgene expression in other cell types such as bone marrow CD34+ progenitor cells, in vitro cultured bone marrow mesenchymal cells and phytohemagglutinin A (PHA)-stimulated peripheral blood T-cells. Transgene expression after electroporation with mRNA encoding the enhanced green fluorescent protein (EGFP) was evaluated by flow cytometry. Flow cytometric analysis 24h after EGFP mRNA electroporation revealed that 35% of fresh uncultured bone marrow-derived CD34+ hematopoietic progenitor cells was efficiently transfected. In the population of in vitro cultured mesenchymal cells, 90% of the viable CD45-CD13+ cells showed detectable EGFP expression. The level of transient EGFP expression in the PHA-stimulated T-cell population was 47% at 24h after electroporation. Importantly, no significant difference of cell mortality between electroporated and non-electroporated cells was observed, due to the mild electroporation procedure ensuring high cell viability. When looking at the kinetics of EGFP expression, we saw that EGFP was still expressed at day 7 after electroporation with highest expression at day 4. Short-term gene introduction by mRNA electroporation in hematopoietic progenitor cells, mesenchymal cells and T-cells could be used to direct the differentiation and/or to modulate the function of these cells. We conclude that mRNA electroporation is an efficient method for short-term gene transfer in CD34+ cells, mesenchymal cells and activated T-cells with potential applications in gene therapy procedures.
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Whyte, Lyle G., and William E. Inniss. "Electroporation and its effect on the psychrotrophic bacterium Bacillus psychrophilus." Canadian Journal of Microbiology 40, no. 2 (February 1, 1994): 83–89. http://dx.doi.org/10.1139/m94-014.
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The psychrotrophic bacterium Bacillus psychrophilus was successfully transformed by electroporation with the cloning vector pC194 and the expression vector pPL708. Optimal electroporation parameters such as field strength, pulse length, and electroporation medium, as well as the influence of the growth phase of the bacterial cells, were determined. Maximum transformation efficiencies of 104 transformants/μg plasmid DNA were achieved with late logarithmic – early stationary phase cells grown in medium supplemented with 33.3 mM glycine and electroporated at high field strengths of 18–20 kV∙cm−1. By means of this procedure, a kanamycin-resistant gene was directly cloned into pPL708 and the recombinant plasmid electrotransformed into B. psychrophilus. In addition, the effect of electroporation on protein synthesis was analyzed by 2-dimensional polyacrylamide gel electrophoresis and computing scanning laser densitometry. At least eight proteins were induced by electroporation field strengths of 18 or 25 kV during the first 2 h immediately following electroporation. Conversely, the synthesis of at least five proteins was repressed by electroporation.Key words: electroporation, psychrotrophic bacterium.
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Sales Conniff, Amanda, Jared Tur, Kristopher Kohena, Min Zhang, Justin Gibbons, and Loree C. Heller. "Transcriptomic Analysis of the Acute Skeletal Muscle Effects after Intramuscular DNA Electroporation Reveals Inflammatory Signaling." Vaccines 10, no. 12 (November 29, 2022): 2037. http://dx.doi.org/10.3390/vaccines10122037.
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Skeletal muscle is a promising tissue for therapeutic gene delivery because it is highly vascularized, accessible, and capable of synthesizing protein for therapies or vaccines. The application of electric pulses (electroporation) enhances plasmid DNA delivery and expression by increasing membrane permeability. Four hours after plasmid electroporation, we evaluated acute gene and protein expression changes in mouse skeletal muscle to identify regulated genes and genetic pathways. RNA sequencing followed by functional annotation was used to evaluate differentially expressed mRNAs. Our data highlighted immune signaling pathways that may influence the effectiveness of DNA electroporation. Cytokine and chemokine protein levels in muscle lysates revealed the upregulation of a subset of inflammatory proteins and confirmed the RNA sequencing analysis. Several regulated DNA-specific pattern recognition receptor mRNAs were also detected. Identifying unique molecular changes in the muscle will facilitate a better understanding of the underlying molecular mechanisms and the development of safety biomarkers and novel strategies to improve skeletal muscle targeted gene therapy.
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Chau, Chalmers, Paolo Actis, and Eric Hewitt. "Methods for protein delivery into cells: from current approaches to future perspectives." Biochemical Society Transactions 48, no. 2 (April 8, 2020): 357–65. http://dx.doi.org/10.1042/bst20190039.
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The manipulation of cultured mammalian cells by the delivery of exogenous macromolecules is one of the cornerstones of experimental cell biology. Although the transfection of cells with DNA expressions constructs that encode proteins is routine and simple to perform, the direct delivery of proteins into cells has many advantages. For example, proteins can be chemically modified, assembled into defined complexes and subject to biophysical analyses prior to their delivery into cells. Here, we review new approaches to the injection and electroporation of proteins into cultured cells. In particular, we focus on how recent developments in nanoscale injection probes and localized electroporation devices enable proteins to be delivered whilst minimizing cellular damage. Moreover, we discuss how nanopore sensing may ultimately enable the quantification of protein delivery at single-molecule resolution.
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Campillo-Davo, Diana, Maxime De Laere, Gils Roex, Maarten Versteven, Donovan Flumens, Zwi N. Berneman, Viggo F. I. Van Tendeloo, Sébastien Anguille, and Eva Lion. "The Ins and Outs of Messenger RNA Electroporation for Physical Gene Delivery in Immune Cell-Based Therapy." Pharmaceutics 13, no. 3 (March 16, 2021): 396. http://dx.doi.org/10.3390/pharmaceutics13030396.
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Messenger RNA (mRNA) electroporation is a powerful tool for transient genetic modification of cells. This non-viral method of genetic engineering has been widely used in immunotherapy. Electroporation allows fine-tuning of transfection protocols for each cell type as well as introduction of multiple protein-coding mRNAs at once. As a pioneering group in mRNA electroporation, in this review, we provide an expert overview of the ins and outs of mRNA electroporation, discussing the different parameters involved in mRNA electroporation as well as the production of research-grade and production and application of clinical-grade mRNA for gene transfer in the context of cell-based immunotherapies.
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Bertling, Wolf. "Transfection of a DNA/protein complex into nuclei of mammalian cells using polyoma capsids and electroporation." Bioscience Reports 7, no. 2 (February 1, 1987): 107–12. http://dx.doi.org/10.1007/bf01121873.
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We used empty capsids ofpolyoma virus to transfer DNA fragments and DNA/protein complexes into human cells. We encapsulated labeled and unlabeled single stranded DNA fragments by viral capsids. A complex of DNA with a DNA binding protein, recA, will also be taken up by the capsids, whereas the free protein is not incorporated. We further compared this gentle biological method of DNA transfection with a well-established physical method, electroporation. Electroporation also allows the transfer of DNA as well as protein into cells, although there is no proof that a DNA/protein complex can survive the procedure functionally. Whereas the viability of capsid transfected cells is unaffected (100%), electroporation reduces the viability to 90–95%. On the other hand, the amount of DNA found in the nucleus of electroporated cells is higher than for cells treated with loaded viral capsids.
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Dai, Yang, Yinchang Zhu, Donald A. Harn, Xiaoting Wang, Jianxia Tang, Song Zhao, Fei Lu, and Xiaohong Guan. "DNA Vaccination by Electroporation and Boosting with Recombinant Proteins Enhances the Efficacy of DNA Vaccines for Schistosomiasis Japonica." Clinical and Vaccine Immunology 16, no. 12 (October 7, 2009): 1796–803. http://dx.doi.org/10.1128/cvi.00231-09.
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ABSTRACT Schistosomiasis japonica is an endemic, zoonotic disease of major public health importance in China. Control programs combining chemotherapy and snail killing have not been able to block transmission of infection in lakes and marsh regions. Vaccination is needed as a complementary approach to the ongoing control programs. In the present study, we wanted to determine if the efficacies of DNA vaccines encoding the 23-kDa tetraspanin membrane protein (SjC23), triose phosphate isomerase (SjCTPI), and sixfold-repeated genes of the complementarity determining region 3 (CDR3) in the H chain of NP30 could be enhanced by boosting via electroporation in vivo and/or with cocktail protein vaccines. Mice vaccinated with cocktail DNA vaccines showed a significant worm reduction of 32.88% (P < 0.01) and egg reduction of 36.20% (P < 0.01). Vaccine efficacy was enhanced when animals were boosted with cocktail protein vaccines; adult worm and liver egg burdens were reduced 45.35% and 48.54%, respectively. Nearly identical results were obtained in mice boosted by electroporation in vivo, with adult worm and egg burdens reduced by 45.00% and 50.88%, respectively. The addition of a protein vaccine boost to this regimen further elevated efficacy to approximately 60% for adult worm burden and greater than 60% for liver egg reduction. The levels of interleukin-2, gamma interferon, and the ratios of immunoglobulin G2a (IgG2a)/IgG1 clearly showed that cocktail DNA vaccines induced CD4+ Th1-type responses. Boosting via either electroporation or with recombinant proteins significantly increased associated immune responses over those seen in mice vaccinated solely with DNA vaccines. Thus, schistosome DNA vaccine efficacy was significantly enhanced via boosting by electroporation in vivo and/or cocktail protein vaccines.
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Muriel, Joaquin M., Andrea O’Neill, Jaclyn P. Kerr, Emily Kleinhans-Welte, Richard M. Lovering, and Robert J. Bloch. "Keratin 18 is an integral part of the intermediate filament network in murine skeletal muscle." American Journal of Physiology-Cell Physiology 318, no. 1 (January 1, 2020): C215—C224. http://dx.doi.org/10.1152/ajpcell.00279.2019.
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Intermediate filaments (IFs) contribute to force transmission, cellular integrity, and signaling in skeletal muscle. We previously identified keratin 19 (Krt19) as a muscle IF protein. We now report the presence of a second type I muscle keratin, Krt18. Krt18 mRNA levels are about half those for Krt19 and only 1:1,000th those for desmin; the protein was nevertheless detectable in immunoblots. Muscle function, measured by maximal isometric force in vivo, was moderately compromised in Krt18-knockout ( Krt18-KO) or dominant-negative mutant mice ( Krt18 DN), but structure was unaltered. Exogenous Krt18, introduced by electroporation, was localized in a reticulum around the contractile apparatus in wild-type muscle and to a lesser extent in muscle lacking Krt19 or desmin or both proteins. Exogenous Krt19, which was either reticular or aggregated in controls, became reticular more frequently in Krt19-null than in Krt18-null, desmin-null, or double-null muscles. Desmin was assembled into the reticulum normally in all genotypes. Notably, all three IF proteins appeared in overlapping reticular structures. We assessed the effect of Krt18 on susceptibility to injury in vivo by electroporating siRNA into tibialis anterior (TA) muscles of control and Krt19-KO mice and testing 2 wk later. Results showed a 33% strength deficit (reduction in maximal torque after injury) compared with siRNA-treated controls. Conversely, electroporation of siRNA to Krt19 into Krt18-null TA yielded a strength deficit of 18% after injury compared with controls. Our results suggest that Krt18 plays a complementary role to Krt19 in skeletal muscle in both assembling keratin-based filaments and transducing contractile force.
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Lan, Chen-Yu, Ping-Heng Tan, Jiin-Tsuey Cheng, Hsiao-Feng Lu, Ming-Wei Lin, Po-Ni Hsiao, and Chung-Ren Lin. "Immunoneutralization of c-Fos Using Intrathecal Antibody Electroporation Attenuates Chronic Constrictive Injury-induced Hyperalgesia and Regulates Preprodynorphin Expression in Rats." Anesthesiology 99, no. 4 (October 1, 2003): 938–46. http://dx.doi.org/10.1097/00000542-200310000-00029.
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Background In vivo electroporation has been successfully used for the introduction of DNA, RNA, oligonucleotides, and proteins into cells for experimental and therapeutic purposes. The authors evaluated the efficacy of electroporation-mediated c-Fos antibody therapy for neuropathic pain in vitro and in vivo. Methods First, the authors studied the inhibitory effects of intrathecal c-Fos antibody electroporation on the activating protein (AP-1) promoter activity in cultured spinal neuronal cells transfected with p-AP-Luc plasmid and activated with 100 microm glutamate. The inhibitory effect of c-Fos antibody electroporation in the regulation of AP-1 promoter activity was assessed according to the relative luciferase activity. Second, rats with chronic constrictive injury underwent electroporation treatment for neuropathic pain using c-Fos antibody. Thermal nociceptive thresholds were measured before chronic constrictive injury and then on even-numbered days, up to and including day 14, to assess and compare the therapeutic effects of intrathecal electroporation. The time course was assessed by Western blot analysis and by immunohistochemical analysis. Pronociceptive gene expression was measured by assessing prodynorphin mRNA and dynorphin peptides on days 2 and 10 after intrathecal c-Fos electroporation. Results Cotransfection of c-Fos antibody significantly decreased glutamate-induced AP-1 activity. Intrathecal electrotransfer of c-Fos antibody attenuated spinal dynorphin levels, as manifested by significantly elevated pain thresholds in the chronic constrictive injury-affected limbs. Conclusion This study shows that transfer of antibody into rat spinal cords by intrathecal electroporation is a useful method to study the function of endogenous factors of spinal-related disorders.
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Lambert, Helene, Roumen Pankov, Johanne Gauthier, and Ronald Hancock. "Electroporation-mediated uptake of proteins into mammalian cells." Biochemistry and Cell Biology 68, no. 4 (April 1, 1990): 729–34. http://dx.doi.org/10.1139/o90-105.
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Proteins of up to 230 kilodaltons are taken up by Chinese hamster ovary fibroblasts exposed to electroporation under conditions generally similar to those used to mediate DNA transfection. The uptake of catalase, ovalbumin, and histone H1 labelled with fluorescein was visualized by fluorescence microscopy. Under the same conditions, the uptake of colloidal gold particles (20 nm diameter) was visualized by electron microscopy. In optimum conditions, about 25% of the cells remained viable and grew normally and about 25% of these retained labelled proteins during two cycles of further growth. About 6 × 104 molecules of catalase were retained per cell. Proteins were taken up when presented to the cells up to 4 h after electroporation, suggesting that mechanisms other than classical electropore formation may operate in these conditions. The proteins were localized in the cytoplasm in a predominantly vesicular pattern and histone H1 entered the nucleus in some cells.Key words: electroporation, protein uptake, gold particle uptake, endocytosis, cell membrane.
Dissertations / Theses on the topic "Protein electroporation"
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Chen, Zhiqiang. "NANOMETER-SCALE MEMBRANE ELECTRODE SYSTEMS FOR ACTIVE PROTEIN SEPARATION, ENZYME IMMOBILIZATION AND CELLULAR ELECTROPORATION." UKnowledge, 2014. http://uknowledge.uky.edu/cme_etds/33.
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Automated and continuous processes are the future trends in downstream protein purification. A functionalized nanometer-scale membrane electrode system, mimicking the function of cell wall transporters, can selectively capture genetically modified proteins and subsequently pump them through the system under programmed voltage pulses. Numerical study of the two-step pulse pumping cycles coupled with experimental His-GFP releasing study reveals the optimal 14s/1s pumping/repel pulse pumping condition at 10 mM bulk imidazole concentration in the permeate side. A separation factor for GFP: BSA of 9.7 was achieved with observed GFP electrophoretic mobility of 3.1×10-6 cm2 s-1 V-1 at 10 mM bulk imidazole concentration and 14 s/1 s pumping/repel duration. The purification of His6-OleD Loki variant directly from crude E. coli extracts expression broth was demonstrated using the pulse pumping process, simplifying the separation process as well as reducing biopharmaceutical production costs. The enzymatic reactions showed that His6-OleD Loki was still active after purification.
A nanoporous membrane/electrode system with directed flow carrying reagents to sequentially attached enzymes to mimic nature’s enzymes-complex system was demonstrated. The substrates residence time on the immobilized enzyme can be precisely controlled by changing the pumping rate and thereby prevent a secondary hydrolysis reaction. Immobilized enzyme showed long term storage longevity with activity half-life of 50 days at 4℃ and the ability to be regenerated. One-step immobilization and purification of His-tagged OleD Loki variant directly from expression broth, yielded 98% Uridine Diphosphate glycosylation and 80% 4-methylumbelliferone glycosylation conversion efficiency for the sequential reaction.
A flow-through electroporation system, based on a novel membrane/electrode design, for the delivery of membrane-impermeant molecules into Model Leukocyte cells was demonstrated. The ability to apply low voltage between two short distance electrodes contributes to high cell viability. The flow-through system can be easily scaled-up by varying the micro-fluidic channel geometry and/or the applied voltage pulse frequency. More importantly, the system allows the electrophoretical pumping of molecules from the reservoir across the membrane/electrode system to the micro-fluidic channel for transfection, which reduces large amount of reagents used.
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Schmotzer, Carolyn Anne. "Assessment of Murine Embryo Development Following Electroporation and Microinjection of a Green Fluorescent Protein DNA Construct." Thesis, Virginia Tech, 2001. http://hdl.handle.net/10919/34369.
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Transgenic techniques have rapidly evolved in recent years. However, the efficiency of these techniques to produce viable offspring is still disappointingly low. The purpose of this study was to assess in vitro development, transgene expression, and integration following pronuclear or cytoplasmic microinjection of condensed or linear green fluorescent protein DNA into murine embryos using electroporation. In experiment 1, the effect of embryo orientation (group or linear) within the electroporation chamber on development was evaluated using zygotes which received one pulse duration (10 msec), and one of two voltages (250 or 400 V). Zygotes that received 400 V had the lowest development score (Group, 2.06 ? 0.12; Linear, 1.97 ? 0.13), irrespective of orientation. Embryos that received 250 V had the highest development of the voltage treated groups (Group 3.42 ? 0.12; Linear 3.32 ? 0.12), irrespective of orientation, and development was lower than the control embryos (Control 4.28 ? 0.12; Mannitol control 4.36 ? 0.18). In experiment 2, the efficiency of utilization of the prepared enhanced green fluorescent protein (EGFP) construct as a visual marker of protein expression was evaluated using pronuclear microinjection. Embryo development and fluorescence were evaluated following pronuclear injection of EGFP at a concentration of 3 μg/ml and compared to an uninjected control. Embryos injected with the EGFP had lower development scores (3.85 ± 0.15) than uninjected control embryos (5.72 ± 0.2). Of the embryos injected, 32.4% fluoresced due to expression of EGFP. Experiment 3 evaluated the effect of combining cytoplasmic injection of EGFP (425 μg/ml) with electroporation at 250 V on EGFP expression. The non-manipulated control embryos had significantly higher (P < 0.01) 4 d development scores (5.57 ± 0.11) than manipulated control embryos (4.6 ± 0.18), where the injection needle was inserted into the cytoplasm and no DNA was injected. Combining cytoplasmic DNA injection and electroporation caused a significant (P < 0.01) decrease in development scores, irrespective of DNA construct, when compared to embryos injected with a DNA construct alone. The mechanical effects of needle insertion combined with electroporation were not significantly different (P > 0.05) from embryos injected with DNA alone, irrespective of construct injected. Cytoplasmic injection of condensed DNA (0.38%), linear DNA (0.38%), and condensed DNA combined with electroporation (0.36%) resulted in one fluorescent embryo respectively. Cytoplasmic injection of linear DNA when combined with electroporation (3.57%) resulted in 13 fluorescent embryos. Pronuclear injection of the prepared EGFP construct results in lower development than control embryos. Electrical stimulation of zygotes reduces early embryo development. However, low amounts of electrical stimulation may allow for enhancement of gene integration in transgenic embryos.Master of Science
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Kawai, Mariko. "Ectopic bone formation by human bone morphogenetic protein-2 gene transfer to skeletal muscle using transcutaneous electroporation." Kyoto University, 2004. http://hdl.handle.net/2433/147446.
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McCray, Andrea Nicole. "Electrogenetherapy of established B16 murine melanoma by using an expression plasmid for HIV-1 viral protein R." [Tampa, Fla] : University of South Florida, 2006. http://purl.fcla.edu/usf/dc/et/SFE0001758.
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Hua, Ethan Wei. "Maturation of single retinogeniculate projections visualized by in vivo electroporation of fluorescent proteins." Diss., Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 2008. http://wwwlib.umi.com/cr/ucsd/fullcit?p1459290.
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Thesis (M.S.)--University of California, San Diego, 2008.Title from first page of PDF file (viewed Nov. 10, 2008). Available via ProQuest Digital Dissertations. Includes bibliographical references.
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Figueiredo, Lilybeth de Andrade. "Vacinas, novas perspectivas." Master's thesis, [s.n.], 2014. http://hdl.handle.net/10284/4876.
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Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências FarmacêuticasAs vacinas são uma das maiores descobertas da medicina moderna e têm contribuído para salvar a vida de milhares de milhões de pessoas em cooperação com outras medidas de saúde pública (ao nível do saneamento básico, antibióticos, etc.). As vacinas tiveram um forte impacto no combate a muitas doenças, tendo sido a erradicação da varíola uma das maiores conquistas. No entanto, esta área enfrenta ainda desafios complexos, como são os casos do HIV, tuberculose e malária. Os benefícios das vacinas são elevados quando as mesmas são usadas amplamente, no entanto, os custos de produção, distribuição e preservação são elevados sendo dos principais problemas para os países em desenvolvimento, constituindo desta forma um dos principais obstáculos para alcançar uma cobertura vacinal global. Estas limitações levam à necessidade de torná-las mais eficazes, seguras, de produção mais rápida e eficientes, procurando evitar alguns dos seus maiores problemas como a refrigeração, doses múltiplas e injecções intramusculares. Nas últimas décadas foram pesquisadas e estudadas exaustivamente novas tecnologias associadas às vacinas, assim como, foram optimizadas novas formas de administrar e de apresentar os antigénios aos diversos componentes do sistema imunitário. Estas tecnologias incluem mecanismos de produção em que vacinas produzidas em culturas de células são mais rentáveis em comparação com as vacinas que recorrem a ovos. No entanto têm sido implementadas novas estratégias com o objectivo de aumentar a eficácia das vacinas, têm sido implementadas, como é o caso dos adjuvantes, da eletroporação, das vias de imunização e do prime-boost. Finalmente, tem-se tentado encontrar formas diferentes de activar o sistema imunitário através de; vacinas baseadas em proteínas recombinantes; vacinas de DNA; partículas semelhantes a vírus; vacinas universais; vacinas baseadas em vectores virais e vacinas baseadas em péptidos. Nesta área estão constantemente a surgir novas tecnologias de vacinas mais seguras eficazes e de baixo custo. Apesar de algumas estarem ainda em fase experimental, existe um enorme potencial para o surgimento de novas vacinas num futuro próximo, com produção em larga escala, mais eficazes e seguras. Vaccines are one of the greatest discoveries of modern medicine, they have saved the lives of billions of people in cooperation with other public health measures (sanitation, antibiotics, etc.). The vaccines had an impact in reducing many diseases, being the eradication of smallpox one of it's greatest achievements. However this area still faces difficult challenges, such as HIV, tuberculosis and malaria. The benefits of vaccines are high when used widely, however, the costs of manufacturing, distribution and preservation of vaccines are costly and major impediments to developing countries themselves, and represents a major obstacle to achieving global immunization coverage. These limitations lead to the need of improving vaccines to be more effective, safer, faster and more efficient production thus avoiding some of their biggest problems, such as refrigeration, multiple doses and intramuscular injections. In recent decades there have been exhaustively studied and researched new technologies associated with vaccines, as well as new forms were optimised to manage and present the antigens to the various components of the immune system. These technologies include mechanisms that vaccines produced in cell culture are more cost effective in comparison with the vaccine produced in eggs. Strategies for the purpose of increasing the effectiveness of vaccines such as adjuvants, electroporation, routes of immunisation, and the prime-boost. Finally, find different ways to activate the immune system such as vaccines based on recombinant proteins; DNA vaccines; virus-like particles; universal vaccines; vaccines based on viral vectors and vaccines based on peptide. In this area, there are constantly arising new technologies of safer vaccines,effective and inexpensive. Although some of the technologies are still in experimental stages, there is huge potential for the emergence of new vaccines, with large-scale production, effective and safe in the near future.
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Grognot, Marianne. "Imagerie térahertz par réflexion interne totale pour la biologie. : Application à l'étude de la perméabilisation cellulaire." Thesis, Université Paris-Saclay (ComUE), 2016. http://www.theses.fr/2016SACLX068/document.
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Les ondes térahertz s’étendent de 0.1 à 10x1012 Hz, à la frontière entre les domaines de l’optique et des radiofréquences. Cette position intermédiaire originale en a longtemps rendu l’accès difficile : les technologies térahertz n’ont pris leur essor qu’au cours des années 90. Le domaine n’a pas encore atteint la maturité des domaines des microondes ou de l’infrarouge qui le jouxtent. Cependant, les motivations exploratoires sont fortes, de par la sensibilité spectroscopique du térahertz aux états moléculaires (rotationnels, vibrationnels..) et aux liaisons faibles établies dans et entre les molécules. Dans le cas des objets biologiques, le térahertz est particulièrement sensible à l’eau : sa quantité, son état physico-chimique et ses solutés.Nous avons mis en œuvre un montage d’imagerie en réflexion interne totale atténuée (ATR) pour pouvoir distinguer des cellules vivantes de leur milieu physiologique. Au cours de ce travail, le montage d’imagerie ATR a été caractérisé théoriquement, puis expérimentalement. La première démonstration de l’origine du contraste sur ces images térahertz a été réalisée. Il provient du contenu intracellulaire, plus spécifiquement des protéines et peptides dissouts dans le cytoplasme.Une analyse fine des mécanismes sous-jacents à la nature protéique du contraste térahertz a également été développée. Elle donne accès à des informations spectroscopiques inédites sur l’eau, les protéines dissoutes et la couche de solvatation les entourant.Mettant à profit cette compréhension de notre montage térahertz, nous l’avons proposé comme outil non invasif de suivi quantitatif de la perméabilisation de cellules en conditions physiologiques. Lors de la perméabilisation, augmentation des transferts moléculaires à travers la membrane, notre outil permet de quantifier le passage des peptides et protéines. La perméabilisation de cellules vivantes a une gamme d’application vaste, de l’entrée de fluorochromes pour l’imagerie ou de médicaments à la thérapie génique. Afin d’assurer ces passages à travers la membrane des cellules, il est nécessaire d’altérer ses propriétés, sans pour autant compromettre la viabilité cellulaire. L’étude de deux types de perméabilisation avec notre outil térahertz est proposée : la perméabilisation chimique et l’électroporation. Dans les deux cas, des mécanismes d’effet dose ont été caractérisés quantitativement. Notre outil térahertz a démontré de grands avantages devant les méthodes actuellement utilisées pour quantifier ces dynamiques de perméabilisation et en caractériser la réversibilitéLying between 0.1 to 10x1012 Hz, the terahertz radiation occupies a middle ground between microwaves and infrared light waves, sometimes named “the terahertz gap” for technologies relevant to generation and detection have only risen at the beginning of the 90’s and aren’t fully developed yet. Nevertheless, there are strong exploratory incentives because of terahertz spectroscopic sensitivity to molecular states (rotational, vibrational…) and weak bounds in and between molecules. In the case of biological object, terahertz waves are especially sensitive to water: its quantity, physico-chemical state and solutes. We implemented an Attenuated Total internal Reflection (ATR) imaging setup in order to distinguish live cells from their physiological bathing medium. Throughout this work, we characterized both experimentally and experimentally the ATR setup. The first demonstration of the contrast origin in the terahertz images obtained was done. It arises from the intracellular content, more specifically the proteins and peptides dissolved in the cytoplasm.A precise analysis of the underlying mechanism of this proteinaceous terahertz contrast has also been developed. It gives access to original spectroscopic information about water, dissolved proteins and the hydration shell around them.Taking advantage of our whole setup comprehension, we proposed it as a non-invasive tool for quantitative live-cell permeabilization assessment in physiological conditions. During permeabilization, aka increased molecular transfers through the cell membrane, our tool allows to quantify the transfer of peptides and proteins. Live-cell permeabilization has a large application range, from fluorochrome entry in imaging, to drugs or gene therapy. In order to ensure molecules crossing the cell membrane, it’s necessary to alter its properties without compromising cell viability.A study of two permeabilization methods is proposed: chemical permeabilization and electroporation. In both cases dose effect mechanisms were quantitatively characterized. Our terahertz tool demonstrated great advantages over classical permeabilization quantification methods and permeabilization reversibility assessment methods
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Ferraro, Bernadette. "Intradermal Delivery of Plasmids Encoding Angiogenic Growth Factors by Electroporation Promotes Wound Healing and Neovascularization." [Tampa, Fla] : University of South Florida, 2009. http://purl.fcla.edu/usf/dc/et/SFE0002823.
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Walker, Tara L. "The Development Of Microalgae As A Bioreactor System For The Production Of Recombinant Proteins." Thesis, Queensland University of Technology, 2004. https://eprints.qut.edu.au/15905/1/Tara_Walker_Thesis.pdf.
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Dunaliella, a genus of unicellular, biflagellate green algae, is one of the most studied microalgae for mass culture and is of commercial importance as a source of natural -carotene. Dunaliella species have the desirable properties of halotolerance and photoautotrophy that makes their large-scale culture simple and cheap using resources unsuitable for conventional agriculture. The ease and cost-effectiveness of culture makes Dunaliella a desirable target for increased production of natural compounds by metabolic engineering or for exploitation as biological factories for the synthesis of novel high-value compounds. However, the lack of efficient genetic transformation systems has been a major limitation in the manipulation of these microalgae.
In chapter four we describe the development of a nuclear transformation system for Dunaliella tertiolecta. The gene encoding the phleomycin-binding protein from Streptoalloteichus hindustanus, was chosen as the selectable marker as this protein retains activity at high salt concentrations. To drive expression of the chosen selectable marker, two highly expressed Dunaliella tertiolecta RbcS genes and their associated 5' and 3' regulatory regions were isolated and characterised (chapter three). Dunaliella transformation cassettes containing the RbcS promoter and terminator regions flanking the ble antibiotic resistance gene were constructed. These expression cassettes were tested in Chlamydomonas reinhardtii cells and found to drive expression of the ble gene in this heterologous system. This study also demonstrated that truncation of both the D. tertiolecta RbcS1 and RbcS2 regulatory regions significantly increases the expression of the ble gene in C. reinhardtii cells. To determine if the foreign DNA could stably integrate into the Dunaliella genome, four transformation methods: microprojectile bombardment, glass bead-mediated transformation, PEG-mediated transformation and electroporation were tested and a number of parameters varied. Southern blot analysis revealed that the plasmid DNA transiently entered the Dunaliella cells following electroporation but was rapidly degraded. Following electroporation, one stably transformed Dunaliella line was recovered. This is the first demonstration of the stable transformation of this alga.
Chloroplast transformation is becoming a favoured method for the production of recombinant proteins in plants, as levels of heterologous protein are often higher than those achieved by transforming the nucleus. The Dunaliella chloroplast genome has not been genetically characterised, and thus there were no existing promoter and terminator sequences or sequences of intergenic regions that could be used for vectors in transformation of the chloroplast. Therefore, this study aimed to isolate and characterise promoters of highly expressed genes and matching terminators capable of driving transgene expression, and also to characterise intergenic regions that would be suitable insertion sites for the vector construct (chapter five). The complete gene sequence of two highly expressed Dunaliella chloroplast genes psbB and rbcL including the promoter and terminator regions as well as the coding sequence of the psbA gene were cloned and sequenced. In addition, the psbA gene is useful as a selectable marker as introduced mutations confer resistance to the herbicide 3-(3,4-Dichlorophenyl)-1,1-Dimethylurea (DCMU). Two homologous transformation constructs based on mutated psbA genes were developed and tested using microprojectile bombardment. A number of parameters were tested including: the size of the gold microprojectile particle, the distance of the plates from the point of discharge, plating onto membranes or filter paper, helium pressure, addition of an osmoticum to the medium and recovery time. Although no chloroplast transformants were recovered in this study, these homologous recombination constructs should prove useful in the development of a chloroplast transformation protocol.
The other major component of this study was to investigate the use of microalgae as an expression system for the production of recombinant proteins. Transformation of Chlamydomonas reinhardtii, a species related to Dunaliella, is well developed. In chapter six, this study examined the expression of two human proteins, -lactalbumin and IGF-1 in Chlamydomonas reinhardtii. Plasmids containing the C. reinhardtii RbcS2 promoter upstream of the cDNAs of these two proteins were introduced into C. reinhardtii cells using glass-bead mediated transformation. Transgenic C. reinhardtii lines were generated and shown to contain the transgenes by PCR and Southern hybridisation. RT- PCR and northern hybridisation were subsequently used to demonstrate that the transgenes were transcriptionally active. The transcripts however, could only be detected by RT-PCR indicating that the genes were transcribed at low levels. Accumulation of the -lactalbumin protein could not be demonstrated, suggesting that although the transgenes were transcribed, they were either not translated or translated at levels below the sensitivity of western blot analysis or that any protein produced was rapidly degraded. Previous studies have indicated that in microalgae codon usage is vital in translation of the foreign protein. Codon modification of the IGF-I and -lactalbumin genes should lead to higher levels of protein accumulation.
This study reports the first successful stable nuclear transformation of Dunaliella tertiolecta. Therefore it is now feasible that Dunaliella can be examined as a bioreactor for the expression of recombinant proteins. In addition, two chloroplast genes (psbB and rbcL) and their corresponding promoters and terminators have been characterised and a selectable marker cassette based on the mutated psbA gene constructed.
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Walker, Tara L. "The Development Of Microalgae As A Bioreactor System For The Production Of Recombinant Proteins." Queensland University of Technology, 2004. http://eprints.qut.edu.au/15905/.
Full textAbstract:
Dunaliella, a genus of unicellular, biflagellate green algae, is one of the most studied microalgae for mass culture and is of commercial importance as a source of natural -carotene. Dunaliella species have the desirable properties of halotolerance and photoautotrophy that makes their large-scale culture simple and cheap using resources unsuitable for conventional agriculture. The ease and cost-effectiveness of culture makes Dunaliella a desirable target for increased production of natural compounds by metabolic engineering or for exploitation as biological factories for the synthesis of novel high-value compounds. However, the lack of efficient genetic transformation systems has been a major limitation in the manipulation of these microalgae. In chapter four we describe the development of a nuclear transformation system for Dunaliella tertiolecta. The gene encoding the phleomycin-binding protein from Streptoalloteichus hindustanus, was chosen as the selectable marker as this protein retains activity at high salt concentrations. To drive expression of the chosen selectable marker, two highly expressed Dunaliella tertiolecta RbcS genes and their associated 5' and 3' regulatory regions were isolated and characterised (chapter three). Dunaliella transformation cassettes containing the RbcS promoter and terminator regions flanking the ble antibiotic resistance gene were constructed. These expression cassettes were tested in Chlamydomonas reinhardtii cells and found to drive expression of the ble gene in this heterologous system. This study also demonstrated that truncation of both the D. tertiolecta RbcS1 and RbcS2 regulatory regions significantly increases the expression of the ble gene in C. reinhardtii cells. To determine if the foreign DNA could stably integrate into the Dunaliella genome, four transformation methods: microprojectile bombardment, glass bead-mediated transformation, PEG-mediated transformation and electroporation were tested and a number of parameters varied. Southern blot analysis revealed that the plasmid DNA transiently entered the Dunaliella cells following electroporation but was rapidly degraded. Following electroporation, one stably transformed Dunaliella line was recovered. This is the first demonstration of the stable transformation of this alga. Chloroplast transformation is becoming a favoured method for the production of recombinant proteins in plants, as levels of heterologous protein are often higher than those achieved by transforming the nucleus. The Dunaliella chloroplast genome has not been genetically characterised, and thus there were no existing promoter and terminator sequences or sequences of intergenic regions that could be used for vectors in transformation of the chloroplast. Therefore, this study aimed to isolate and characterise promoters of highly expressed genes and matching terminators capable of driving transgene expression, and also to characterise intergenic regions that would be suitable insertion sites for the vector construct (chapter five). The complete gene sequence of two highly expressed Dunaliella chloroplast genes psbB and rbcL including the promoter and terminator regions as well as the coding sequence of the psbA gene were cloned and sequenced. In addition, the psbA gene is useful as a selectable marker as introduced mutations confer resistance to the herbicide 3-(3,4-Dichlorophenyl)-1,1-Dimethylurea (DCMU). Two homologous transformation constructs based on mutated psbA genes were developed and tested using microprojectile bombardment. A number of parameters were tested including: the size of the gold microprojectile particle, the distance of the plates from the point of discharge, plating onto membranes or filter paper, helium pressure, addition of an osmoticum to the medium and recovery time. Although no chloroplast transformants were recovered in this study, these homologous recombination constructs should prove useful in the development of a chloroplast transformation protocol. The other major component of this study was to investigate the use of microalgae as an expression system for the production of recombinant proteins. Transformation of Chlamydomonas reinhardtii, a species related to Dunaliella, is well developed. In chapter six, this study examined the expression of two human proteins, -lactalbumin and IGF-1 in Chlamydomonas reinhardtii. Plasmids containing the C. reinhardtii RbcS2 promoter upstream of the cDNAs of these two proteins were introduced into C. reinhardtii cells using glass-bead mediated transformation. Transgenic C. reinhardtii lines were generated and shown to contain the transgenes by PCR and Southern hybridisation. RT- PCR and northern hybridisation were subsequently used to demonstrate that the transgenes were transcriptionally active. The transcripts however, could only be detected by RT-PCR indicating that the genes were transcribed at low levels. Accumulation of the -lactalbumin protein could not be demonstrated, suggesting that although the transgenes were transcribed, they were either not translated or translated at levels below the sensitivity of western blot analysis or that any protein produced was rapidly degraded. Previous studies have indicated that in microalgae codon usage is vital in translation of the foreign protein. Codon modification of the IGF-I and -lactalbumin genes should lead to higher levels of protein accumulation. This study reports the first successful stable nuclear transformation of Dunaliella tertiolecta. Therefore it is now feasible that Dunaliella can be examined as a bioreactor for the expression of recombinant proteins. In addition, two chloroplast genes (psbB and rbcL) and their corresponding promoters and terminators have been characterised and a selectable marker cassette based on the mutated psbA gene constructed.
Book chapters on the topic "Protein electroporation"
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Yu, Xiaoxi, Olivier Bals, Nabil Grimi, and Eugène Vorobiev. "Polyphenol and Protein Extraction from Rapeseed Stems and Leaves Assisted by Pulsed Electric Fields." In Handbook of Electroporation, 1–17. Cham: Springer International Publishing, 2016. http://dx.doi.org/10.1007/978-3-319-26779-1_122-1.
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Yu, Xiaoxi, Olivier Bals, Nabil Grimi, and Eugène Vorobiev. "Polyphenol and Protein Extraction from Rapeseed Stems and Leaves Assisted by Pulsed Electric Fields." In Handbook of Electroporation, 2733–49. Cham: Springer International Publishing, 2017. http://dx.doi.org/10.1007/978-3-319-32886-7_122.
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Laustsen, Anders, and Rasmus O. Bak. "Electroporation-Based CRISPR/Cas9 Gene Editing Using Cas9 Protein and Chemically Modified sgRNAs." In Methods in Molecular Biology, 127–34. New York, NY: Springer New York, 2019. http://dx.doi.org/10.1007/978-1-4939-9170-9_9.
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Meglič, S. Haberl, E. Levičnik, E. Luengo, J. Raso, and D. Miklavčič. "The Effect of Temperature on Protein Extraction by Electroporation and on Bacterial Viability." In 1st World Congress on Electroporation and Pulsed Electric Fields in Biology, Medicine and Food & Environmental Technologies, 175–78. Singapore: Springer Singapore, 2016. http://dx.doi.org/10.1007/978-981-287-817-5_39.
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Weaver, James C. "Membrane Electroconformational Changes: Progress in Theoretical Modelling of Electroporation and of Protein Protrusion Alteration." In Charge and Field Effects in Biosystems—3, 477–96. Boston, MA: Birkhäuser Boston, 1992. http://dx.doi.org/10.1007/978-1-4615-9837-4_38.
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Coustets, Mathilde, and Justin Teissié. "The Use of Pulsed Electric Fields for Protein Extraction from Nanochloropsis and Chlorella." In 1st World Congress on Electroporation and Pulsed Electric Fields in Biology, Medicine and Food & Environmental Technologies, 405–8. Singapore: Springer Singapore, 2016. http://dx.doi.org/10.1007/978-981-287-817-5_88.
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Fujishiro, Kensei, Yuka Fukui, Osamu Sato, Kohei Kawabe, Koichi Seto, and Kiyoto Motojima. "Analysis of tissue-specific and PPARα-dependent induction of FABP gene expression in the mouse liver by an in vivo DNA electroporation method." In Cellular Lipid Binding Proteins, 165–72. Boston, MA: Springer US, 2002. http://dx.doi.org/10.1007/978-1-4419-9270-3_21.
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Di Paolo, Diana, and Richard M. Berry. "Imaging of Single Dye-Labeled Chemotaxis Proteins in Live Bacteria Using Electroporation." In Methods in Molecular Biology, 233–46. New York, NY: Springer New York, 2018. http://dx.doi.org/10.1007/978-1-4939-7577-8_19.
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Gerer, Kerstin F., Stefanie Hoyer, Jan Dörrie, and Niels Schaft. "Electroporation of mRNA as Universal Technology Platform to Transfect a Variety of Primary Cells with Antigens and Functional Proteins." In RNA Vaccines, 165–78. New York, NY: Springer New York, 2016. http://dx.doi.org/10.1007/978-1-4939-6481-9_10.
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Hayashi, Masahito, and Ritsu Kamiya. "Protein Electroporation into Chlamydomonas for Mutant Rescue." In Methods in Cell Biology, 107–11. Elsevier, 2009. http://dx.doi.org/10.1016/s0091-679x(08)92007-0.
Full textConference papers on the topic "Protein electroporation"
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Ringwelski, Beth, Vidura Jayasooriya, and Dharmakeerthi Nawarathna. "Label Free Cell Purification Following Electroporation." In 2020 Design of Medical Devices Conference. American Society of Mechanical Engineers, 2020. http://dx.doi.org/10.1115/dmd2020-9037.
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Abstract Cell transfection by electroporation is a biological assay that has been utilized to inject exogenous molecules (e.g.: RNA, DNA and protein) into live cells. Recently, electroporation has been utilized in developing cell therapy for cancer (e.g., CAR T-cell). One of the major drawbacks in current electroporation methods is the cell death during the process. These dead cells can be detrimental, if injected back to the patients. Current cell filtering methods are unable purify T-cells following electroporation, this is due to the lack of unique biomarkers that target the apoptosis and necrosis of T-cells. To address this issue, we have developed a method using dielectrophoresis and microfluidics, where no prior labeling is needed to isolate dead cells from live cells. Upon electroporation, the cell sample has to be flowed through the microfluidic chip where a selective electric field is applied through specially designed electrodes so that the dead cells are trapped on the electrodes, and the live cells are able to flow through and are collected at the end. Results after purification of the cells using our method reveal that it is possible to achieve ∼100% of purity in filtering of the live cells. This method presents a viable solution to a critical concern regarding CAR T-cell manufacturing. This paper presents an extended study of the variation of efficacy in the design with the time from the electroporation.
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Pang, W. K., Y. Zhao, J. Peng, B. Peng, and Y. Xu. "Therapeutic protein production in vivo after electroporation-assisted intramuscular gene delivery." In 2005 IEEE Engineering in Medicine and Biology 27th Annual Conference. IEEE, 2005. http://dx.doi.org/10.1109/iembs.2005.1615919.
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Qin, Zhenpeng, Jing Jiang, Gary Long, and John C. Bischof. "Irreversible Electroporation: An In Vivo Study Within the Dorsal Skin Fold Chamber." In ASME 2011 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2011. http://dx.doi.org/10.1115/sbc2011-53936.
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Electroporation has been traditionally used to enhance molecular transport into cells (e.g. gene therapy) and through tissues (e.g. skin) by creating reversible pores with short electrical pulses [1]. Increasing the parameters (electrical field, pulse duration and number) can induce irreversible damage to the cells and tissue. Recently, irreversible electroporation (IRE) has been investigated as a new tumor ablation method [2]. The advantages of the IRE include the simple and fast procedure (train of μs pulses), sharp demarcation between treated and untreated regions, destruction of tumor cells while preserving the connective tissue, and minimal effect of immune response on treatment efficacy [3]. The unique interaction of electrical field with heterogeneous structures prevents damage to nerves, blood vessels and ducts [4]. IRE has been claimed to produce negligible thermal injury and protein denaturation typical to thermal ablation [5]. However, how each electroporation parameter in IRE affects tumor destruction and the possibility of heating remains to be studied in tumors vivo.
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Mamman, Hassan Buhari, Muhammad Mahadi Abdul Jamil, and Mohamad Nazib Adon. "Studying the influence of electroporation on HT29 cell line interaction with micro-patterned extracellular matrix protein (fibronectin)." In 2016 6th IEEE International Conference on Control System, Computing and Engineering (ICCSCE). IEEE, 2016. http://dx.doi.org/10.1109/iccsce.2016.7893587.
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Trinkle, Christine A., Christopher J. Morgan, and Luke P. Lee. "High Precision Assembly of Soft-Polymer Microfluidic Circuits." In ASME 2006 International Mechanical Engineering Congress and Exposition. ASMEDC, 2006. http://dx.doi.org/10.1115/imece2006-14631.
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Microfluidic chips have made it possible to manipulate biological fluidic samples in increasingly smaller volumes—even enabling multiplexed study of individual cells. Performing biological assays using microfluidic technology not only makes them more portable when compared to their traditional counterparts, but also decreases testing time and cost. These biofluidic circuits vary widely in design and function: multiplexed cell electroporation, on-chip cell culturing, cell-cell communication monitoring, protein crystallization, and small volume sample analysis are only a few examples of potential applications. The rapid rate of growth and change in this field creates a need for inexpensive and flexible rapid prototyping of microfluidic chips.
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Quertermous, T., J. M. Schnee, M. S. Runge, G. R. Matsueda, N. W. Hudson, J. G. Seidman, and E. Haber. "EXPRESSION OF A RECOMBINANT ANTIBODY-TARGETED THROMBOLYTIC MOLECULE." In XIth International Congress on Thrombosis and Haemostasis. Schattauer GmbH, 1987. http://dx.doi.org/10.1055/s-0038-1644616.
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We have recently shown that targeting tissue-type plasminogen activator (t-PA) by covalent linkage to a fibrin-specific monoclonal antibody (59D8) produces a more potent thrombolytic agent which also induces less fibrinogenolysis. A recombinant molecule encoding a t-PA-59D8 fusion protein was constructed to provide a ready source of this agent for further study, and to allow tailoring of the active moities for maximal activity. DNA sequence coding for the 59D8 heavy chain (HC) antigen combining site was cloned from a lambdaphage library by selection with a joining region probe. Gene segments coding for this cloned HC rearrangement, the amino portion of the mouse gamma 2b HC constant region, and the catalytic B chain of t-PA were joined in the pSV2-gpt expression vector. The desired coding sequence was confirmed by nucleotide sequence analysis. The construct was transfected by electroporation into 59D8 hybridoma HC loss variants. Transfectants were screened for antifibrin antibody activity. Positive clones were shown to produce mRNA which hybridized to the human t-PA gene in Northern blot analysis. Supernatants from 5 of these clones were subjected to affinity chromatography on a synthetic fibrin-like peptide-Sepharose column followed by a benzamidine-Sepharose column. Western blots of SDS polyacrylamide gels run under reducing conditions revealed binding to a 60 kd band by a monoclonal antihuman t-PA antibody, consistent with a 59D8 HC-t-PA fusion protein. Also, binding to a 25 kd band by goat anti-mouse Fab indicated the presence of 59D8 light chain. Affinity purified protein was shown to have amidolytic activity of similar potency to t-PA in a chromogenic substrate assay utilizing S-2288. Bifunctionality of the purified protein was demonstrated first by an assay which requires the protein to bind to immobilized fibrin and simultaneously exhibit activity in the S2288 assay, and second by simultaneous fibrin and iodinated anti-t-PA binding.
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Sadik, Mohamed M., Jerry Shan, David Shreiber, and Hao Lin. "Extreme Elongation of Vesicles Under DC Electric Fields." In ASME 2008 Summer Bioengineering Conference. American Society of Mechanical Engineers, 2008. http://dx.doi.org/10.1115/sbc2008-193243.
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Motive . The motivation of the current study is to investigate the response of vesicles to applied electric fields, with potential applications in electroporation-mediated molecular delivery [3]. In this technique, an applied field transiently permeabilizes the cell membrane to gain access to the cytoplasm, and deliver active agents such as genes, RNA, proteins into the cell. Although widely applied in classical and emerging areas such as drug delivery and stem cell research, current electroporation techniques suffer from low efficiency and high cell death [4]. The present work is a step towards understanding the complex fundamental processes involved in electroporation, and possibly improving it via parametric optimization. For this purpose we use vesicular cellular mimics as our model to provide good controllability, and to focus on the dynamics of the lipid membrane. Our preliminary results show extreme elongation of the vesicles under high-strength, short-duration DC pulses. Such deformation may significantly affect electroporation, and hence the efficiency of molecular delivery.
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Agarwal, Pooja, Stephen Schauer, Xiaohong Ma, Jacques Plummer, Tim Chan, Lindsay Williams, Michele Kaloss, et al. "Abstract B127: Pharmacodynamics and functionality of RheoSwitch® regulated immunomodulatory proteins, expressed from a multigenic embedded cellular bioreactor following intramuscular electroporation in mice." In Abstracts: AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics--Oct 19-23, 2013; Boston, MA. American Association for Cancer Research, 2013. http://dx.doi.org/10.1158/1535-7163.targ-13-b127.
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