SBIR-STTR Award

This Project, the Dracula Pipette Project, is intended to improve on a device invented by the Project Director (Dr. Paul Taylor) and to find ways to manufacture the device for general use. The device is a co-axial microinjection tool in which the injection pipette is presented from inside the holding pipette. This allows it to suck and hold a portion of the nearside of the surface of an embryo while an injection pipette is passed for injection, aspiration of fluid contents or biopsy of embryonic cells. The Dracula Pipette has been entirely handmade until now, and making it, especially making the delicate glass tips involved, has been too difficult and time-consuming to allow it to be used effectively by other researchers. The real significance of this device is that it allows control and manipulation of embryos at the hatched blastocyst stage of development, after hatching from the zona pellucida and before implantation in the uterus. After hatching, these embryos have no real structure and cannot be held on one side and penetrated from the opposite side, as is done routinely with embryos still in the zona. The human embryo exists for two to three 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 hatched blastocysts of other species and on embryos at all stages before implantation, it is the only tool on the horizon for freezing and biopsy of human hatched blastocysts. The Small Business entity involved, GeneSearch, Inc., will supply the current best prototype of the device for testing and practical use by a variety of respected embryo researchers in a variety of laboratory settings, working with different kinds of animal models. The consultants will provide information about their experiences with the device and their ideas for improving or changing it for specific purposes. The inventor and the company Design Engineer will work to correct design problems as they are identified and will contract with various 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.

Public Health Relevance:
The improved Dracula Pipette, with disposable plastic tips and micromanipulator control, will be an important addition to the instrumentation in almost all biotech labs dealing with embryos. It will open areas of research in freezing and biopsy of large hatched blastocysts, areas previously ignored because the tool was lacking. Most importantly, it will give human fertility clinicians a way to freeze and biopsy human hatched blastocysts, embryos at the most differentiated stage of development before implantation in the uterus.

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.

Public Health Relevance Statement:


Public Health Relevance:
The Dracula Pipette is a self-contained single tool for holding and puncturing pre-implantation embryos. It replaces complicated and expensive combinations of conventional devices. It will open areas of research in biopsy and freezing of relatively large embryos that are now very difficult to manipulate, research areas previously ignored because the tool was lacking. Most importantly, it will give human fertility clinicians a way to biopsy and freeze these later-stage embryos harmlessly for pre-implantation genetic diagnosis, PGD, at the stage of development which has been identified as resulting in the most accurate DNA analysis.

Project Terms:
Address; Area; Arts; base; Biopsy; Bladder; blastocyst; blastomere structure; Businesses; Bypass; Caliber; capsule (pharmacologic); Cattle; Cell division; cell water; Cells; Certification; Chills; clinical application; Collection; commercialization; Communities; Contractor; Contracts; Defect; design; Development; Devices; Diagnosis; DNA; DNA analysis; Electronics; Embryo; Embryo Research; Embryonic Development; Equipment; Equus caballus; Evaluation; experience; Failure (biologic function); Feedback; Fertility; Fiber Optics; Freezing; Funding; Genetic; Genetic Materials; Glass; Goals; hatching; Herpes zoster disease; Human; implantation; improved; 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; preimplantation; Printing; Procedures; Process; Production; prototype; public health relevance; Puncture procedure; Regulatory Affairs; Research; Research Personnel; Resistance; Retrieval; Rupture; Sampling; seal; Shapes; Shipping; Ships; Side; Solutions; Special Equipment; Specialist; Stage at Diagnosis; Staging; Stem cells; Sterilization; Stretching; Structure; success; sucking; Suction; Support System; Surface; System; systems research; Technology; Testing; Texas; Time; tool; Trademark; Training; Transgenic Mice; Tube; United States National Institutes of Health; Universities; user-friendly; Uterus; Vacuum; Validation; Variant; Vision; Water; Work; Zona Pellucida