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: 8918240
• 负责人: Paul Jon Taylor
• 金额: $ 0.95万
• 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/8918240
• 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; Diagnosis; Electronics; Embryo; Embryo Research; Embryonic Development; Equipment; Equus caballus; Evaluation; Failure; Feedback; Fertility; Fiber Optics; Freezing; Funding; Genetic; 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; Mails; 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; Stage at Diagnosis; 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; 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（申请人提供）：Dracula 移液管是一种同轴胚胎显微注射工具，注射移液管从固定移液管内部伸出。当在它们之间的空间中施加真空时，双嵌套尖端密封球形胚胎的表面。这使得胚胎近侧表面的一部分能够通过真空保持，同时通过玻璃移液管进行注射、抽吸液体内容物或对胚胎细胞进行活检。在该项目开始前不久发明了一种专有的激光钻孔活检探针，它提供了一种新的、更好的方法，可以对处于发育阶段的植入前胚胎进行无害活检，从而为植入前遗传学诊断（PGD）提供更准确的 DNA 信息。美国专利商标局在申请之日起不到一年的时间里就颁发了“吸刺”工艺的专利，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头的专利正在申请中。 2011 年 10 月，该项目被 NIH 指定为高度优先项目。该装置适用于胚胎发育直至植入子宫的所有阶段，具有独特的能力在孵化囊胚发育阶段控制和操作胚胎。孵化是指逃离透明带（在最初几次细胞分裂期间包围卵子和胚胎本身的胶囊）的自然过程。从透明带孵化出来后，这些早期胚胎没有真正的结构，不能像使用传统设备对仍在透明带中的胚胎进行常规操作那样，将其固定在一侧并从另一侧穿透。德古拉移液器已证明它对于胚胎冷冻前减少水量非常有价值。项目总监泰勒博士使用该工具从冷冻/解冻的美洲驼胚胎中产生了第一批活婴儿。路易斯安那州立大学的研究人员最近在利用 Dracula 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对小鼠胚胎所做的研究表明，该系统能够从内细胞团中存活地收集一些干细胞，并注射基因 将材料（干细胞）注入小鼠胚胎的囊胚腔中以生产转基因小鼠，这一过程以前需要特殊设备来冷却透明带，以及昂贵的压电锤来穿透透明带。人类胚胎作为孵化的囊胚可能存在两天以上，并且在植入前的这个分化程度最高的阶段还没有好的工具来对其进行操作。因为它体积大且充满液体，所以人类 孵化的囊胚很难成功冷冻，也很难进行活检。尽管 Dracula 移液器对于其他物种的扩张和孵化的囊胚以及植入前各个阶段的胚胎的使用肯定很重要，但它是即将到来的对 8 细胞阶段以上和植入子宫之前的人类孵化囊胚进行无害活检的唯一工具。德古拉移液器项目将提供当前最好的设备原型 由几位受人尊敬的胚胎研究人员在各种实验室环境中进行测试和实际使用，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体验以及针对特定目的改进或更改该设备的想法的反馈。发明者和技术承包商团队将努力纠正发现的设计问题，并将使用适当的微型技术公司来生产特定的改进原型零件。随着这些评估和升级周期的继续，重点将转向所有部件的实际制造，因此可以制造出坚固耐用、用户友好的设备。 呈现给科学界。这个工具，德古拉移液器，为常规程序提供廉价的胚胎操作，例如冷冻前的水体积减少，并且能够对后期胚胎执行以前非常困难或不可能的程序。 GeneSearch, Inc.（申请人小型企业）将通过相关知识产权的许可将 Dracula 移液器和活检探针商业化。我们将维护一个小型设施，用于创建和测试设备的新原型和变体，并与被许可方协调，为新用户提供指导和实际帮助。