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

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.

National Institute of Health (NIH)
Office of The Director, National Institutes of Health (OD)
Small Business Innovation Research Grants (SBIR) - Phase II (R44)
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Special Emphasis Panel (ZRG1-EMNR-S (11))
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Contreras, Miguel A
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Genesearch, Inc.
United States
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Sebrell, T Andrew; Sidar, Barkan; Bruns, Rachel et al. (2018) Live imaging analysis of human gastric epithelial spheroids reveals spontaneous rupture, rotation and fusion events. Cell Tissue Res 371:293-307