Co-axial Microinjection System for Freezing and Biopsy of Early Embryos

用于早期胚胎冷冻和活检的同轴显微注射系统

• 批准号: 8837716
• 负责人: Paul Jon Taylor
• 金额: $ 59.5万
• 依托单位: GENESEARCH, INC.
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-05 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8837716
• 关键词: Address; Area; Arts; Biopsy; Bladder; Businesses; Bypass; Caliber; Cattle; Cell division; Cells; Certification; Chills; Collection; Communities; Contractor; Contracts; DNA; DNA analysis; Defect; Development; Devices; Electronics; Embryo; Embryo Research; Embryonic Development; Equipment; Equus caballus; Evaluation; Failure; Feedback; Fertility; Fiber Optics; Freezing; Funding; Genetic Materials; Glass; Goals; Health; Herpes zoster disease; Human; Injection of therapeutic agent; Inner Cell Mass; Instruction; Intellectual Property; Laboratories; Lasers; Legal patent; Licensing; Life; Light; Lighting; Liquid substance; Llama; Location; Mammals; Metals; Methods; Microinjections; Micromanipulation; Microscope; Modeling; Molds; Montana; Movement; Mus; Needles; Ovum; Penetration; Phase; Plastics; Printing; Procedures; Process; Production; Puncture procedure; Regulatory Affairs; Research; Research Personnel; Resistance; Retrieval; Rupture; Sampling; Shapes; Shipping; Ships; Side; Solutions; Special Equipment; Specialist; Staging; Stem cells; Sterilization; Stretching; Structure; Suction; Support System; Surface; System; Technology; Testing; Texas; Time; Trademark; Training; Transgenic Mice; Tube; United States National Institutes of Health; Universities; Uterus; Vacuum; Validation; Variant; Vision; Water; Work; Zona Pellucida; base; blastocyst; blastomere structure; capsule; cell water; clinical application; commercialization; design; experience; genetic disorder diagnosis; hatching; implantation; improved; preimplantation; prototype; seal; success; sucking; systems research; tool; user-friendly

DESCRIPTION (provided by applicant): The Dracula Pipette is a co-axial embryo microinjection tool, with the injection pipette presented from inside the holding pipette. Double nested tips seal against the surface of the spherical embryo when a vacuum is applied in the space between them. This allows a portion of the nearside surface of an embryo to be held by vacuum while a glass pipette is passed for injection, aspiration of fluid contents or biopsy of embryonic cells. A proprietary laser-drilled biopsy probe, invented shortly before the beginning of the project, provides a new and better method for harm-free biopsy of pre-implantation embryos at a stage of development when they can provide more accurate DNA information for pre-implantation genetic diagnosis (PGD). The US Patent and Trademark Office issued the patent for the "suck-and-puncture" process in less than a year from the date of application, saying there was no relevant prior art to consider. Patents for the device itself and for our proprietary laser-drilled biopsy probe are pending. In October of 2011 this project was designated as high priority by NIH. This device, useful at all stages of embryo development up to the time of implantation in the uterus, is uniquely capable of control and manipulation of embryos at the hatched blastocyst stage of development. Hatching refers to the natural process of escape from the zona pellucida (the capsule that encloses the ovum and the embryo itself during the first few cell divisions). After hatching from the zona, these early embryos have no real structure and cannot be held on one side and penetrated from the opposite side, as is done routinely on embryos still in the zona, using conventional equipment. The Dracula Pipette has shown that it is valuable for water-volume reduction before freezing of embryos. Dr. Taylor, the Project Director, produced the first live babies from frozen/thawed embryos of llamas using this tool. Researchers at LSU, have recently had breakthrough success in freezing of large (900um) equine embryos with the Dracula system, which has not been possible before. Work done by Dr. Taylor at Montana State University with mouse embryos shows the capability of this system for survivable collection of a few stem cells from the inner cell mass and for injection of genetic material (stem cells) into the blastocoel space of mouse embryos for production of transgenic mice, a procedure that previously required special equipment for chilling of the zona, as well as the expensive piezo hammer for penetration of the zona. The human embryo may exist for more than two days as a hatched blastocyst, and there has been no good tool for manipulating it at this most-differentiated stage before implantation. Because it is large and fluid-filled, the human hatched blastocyst is difficult to freeze successfully and difficult to biopsy. Though the Dracula Pipette will surely be important for use with the expanded and hatched blastocysts of other species and on embryos at all stages before implantation, it is the only tool on the horizon for harm-free biopsy of human hatched blastocysts beyond the 8-cell stage and before implantation in the uterus. The Dracula Pipette Project will supply the current best prototype of the device for testing and practical use by several respected embryo researchers in a variety of laboratory settings, working mainly with bovine and mouse embryo models. The consultants will provide feedback about their experiences with the device and their ideas for improving or changing it for specific purposes. The inventor and a team of technical contractors will work to correct design problems as they are identified and will use appropriate micro-technology firms for production of specific improved prototype parts. As these cycles of evaluation and upgrade continue, emphasis will shift to practical manufacture of all parts, so a robust, user-friendly device can be presented to the scientific community. This tool, the Dracula Pipette, offers inexpensive embryo manipulation for routine procedures like water- volume reduction before freezing, and the capability to perform previously very difficult or impossible procedures on later-stage embryos. GeneSearch, Inc., the applicant Small Business, will commercialize the Dracula Pipette and biopsy probe through licensing of the intellectual property involved. We will maintain a small facility for creation and testing of new prototypes and variants of the device, and for coordinatin with licensees to provide instruction and practical assistance for new users.

描述（由申请人提供）：Dracula移液器是一种同轴胚胎显微注射工具，注射移液器从固定移液器内部呈现。当在它们之间的空间中施加真空时，双嵌套尖端密封抵靠球形胚的表面。这允许胚胎的近侧表面的一部分通过真空保持，同时玻璃移液管通过用于注射、抽吸流体内容物或胚胎细胞活检。在该项目开始前不久发明的专有激光钻孔活检探针提供了一种新的更好的方法，用于在发育阶段对植入前胚胎进行无害活检，此时它们可以为植入前遗传诊断（PGD）提供更准确的DNA信息。美国专利和商标局在申请之日起不到一年的时间里就发布了“抽吸穿刺”过程的专利，称没有相关的现有技术可供考虑。该设备本身和我们专有的激光钻孔活检探针的专利正在申请中。2011年10月，该项目被NIH指定为高度优先项目。该装置可用于胚胎发育的所有阶段直至植入子宫的时间，独特地能够控制和操纵发育的孵化胚泡阶段的胚胎。孵化是指从透明膜（在最初的几次细胞分裂期间包围卵子和胚胎本身的囊）中逃逸的自然过程。在从胚胎中孵化出来后，这些早期胚胎没有真实的结构，不能像常规设备对仍在胚胎中的胚胎所做的那样，在一侧固定并从另一侧穿透。Dracula移液器已经表明，它在冷冻胚胎之前减少水体积是有价值的。项目负责人泰勒博士使用这种工具从冷冻/解冻的美洲驼胚胎中生产出了第一批活婴儿。路易斯安那州立大学的研究人员最近在用德古拉系统冷冻大型（900微米）马胚胎方面取得了突破性的成功，这在以前是不可能的。蒙大拿州立大学的泰勒博士用小鼠胚胎所做的工作表明，该系统能够从内细胞团中存活地收集一些干细胞，并将遗传物质注射到小鼠胚胎中。 在这一过程中，我们将干细胞材料（干细胞）植入小鼠胚胎的囊胚腔空间，以生产转基因小鼠，这一过程以前需要专门的设备来冷却囊胚，以及昂贵的压电锤来穿透囊胚。人类胚胎可以作为孵化的囊胚存在两天以上，并且在植入前的这个最分化阶段没有很好的工具来操纵它。因为它很大，充满了液体，人类 孵化囊胚很难成功冷冻，也很难活检。尽管Dracula移液器对于其他物种的扩张和孵化囊胚以及植入前所有阶段的胚胎的使用肯定是重要的，但它是唯一一种对8细胞期以上和植入子宫前的人类孵化囊胚进行无害活检的工具。德古拉移液器项目将提供目前最好的设备原型， 几位受人尊敬的胚胎研究人员在各种实验室环境中进行了测试和实际使用，主要使用牛和小鼠胚胎模型。顾问将提供有关他们使用该设备的经验的反馈，以及他们为特定目的改进或改变该设备的想法。发明者和一组技术承包商将努力纠正发现的设计问题，并将利用适当的微型技术公司生产具体的改进原型部件。随着这些评估和升级周期的继续，重点将转移到所有部件的实际制造上，因此， 呈现给科学界。这种工具，德古拉移液器，提供了廉价的胚胎操作的常规程序，如水体积减少冷冻前，并能够执行以前非常困难或不可能的程序后期胚胎。GeneSearch，Inc.申请人Small Business将通过相关知识产权的许可使Dracula移液管和活检探针商业化。我们将保留一个小型设施，用于创建和测试新原型和设备变体，并与许可证持有者协调，为新用户提供指导和实际帮助。