This Small Business Innovation Research Phase I project aims at developing and commercializing a cost-effective cell injection system based on nanofountain probe (NFP) technology. The NFP has been recently used to investigate drug delivery and direct cell injection. Benefits of this method include easy integration with existing infrastructure, force control of injection, and the ability to control injection volumes at the picoliter level. Injection of large number of cells is necessary for drug discovery and associated large scale genomic and proteomic studies. Such studies require delivery of drugs, conjugated nanoparticles, DNA, siRNA, and proteins into living cells to study spatial and temporal molecular regulatory mechanisms within the cell. Considering the delicate nature of living cells, such a task is nontrivial. For direct drug delivery, micropipette based injection has been used for many years. However, its viability (cell survival) and lack of automation limit its broad use. In this project, a single cell injection system will be developed based on leveraging probe-cell force control and fluidic handling capabilities of NFPs. The system has the potential for parallel cell injection and ultimately automated operation, which would greatly enhance viability as a workhorse tool for cell injection in research labs and pharmaceutical companies.
The broader impact/commercial potential of this project is the development of a new platform for high throughput single cell drug delivery, allowing for efficient evaluation of drug efficacy and the development of diagnostics and next generation therapeutics. The application of nanotechnology to medicine (termed nanomedicine) has provided numerous emerging opportunities in healthcare, particularly given the increasing demand for in vitro toxicology and diagnostics as a pathway towards "personalized medicine". Drug delivery to individual cells and monitoring of associated pathways is at the core of most research toward in vitro diagnostics and toxicology. Personalized medicine will require the systematic incorporation of genetic information from individuals in optimizing preventive and therapeutic care, and much more efficient biomedical tools. Commercialization of this platform would allow research centers and pharmaceutical companies to have access to state-of-the-art nanotechnology tools in their endeavor toward a patient-centered health care system. Furthermore, the new single cell injection system will find utility in laboratories in universities across the U.S., exposing the next generation of scientists to nanotechnology and its impact on medicine.
Exciting progress in biotechnology research in recent years has shown promise for cell reprogramming and extremely sensitive diagnostics, yet this research requires efficient, precise, and gentle delivery of molecules into cells (transfection), for which a robust tool is currently lacking. Research funded by this Phase I Small Business Innovation Research grant supported the development of a method for precise delivery of molecules into cells that is more efficient, more precise, and less invasive than any other method used today. The technique is based on nanofountain probe (NFP) technology, a nanofabricated chip with multiple parallel microfluidic probes, which was invented and patented by Professor Horacio Espinosa at Northwestern University. The NFP transfection technology utilizes electroporation as the transfection method, whereby an electric potential is applied to a target cell, generating small pores in the cell membrane that can allow transport of biomolecules, proteins, or drugs into the cell. This method enables research at the single-cell level, which reduces research and development costs for both cell growth and transfection agents. During this grant period, the following research tasks were achieved: (1) a robust packaging method was designed and tested to enable NFP electroporation to be compatible with either a micromanipulator or atomic force microscope; (2) theoretical modeling was performed to help elucidate the mechanism of probe-based electroporation; (3) parametric experimental studies of voltage, resistance, and dosage control were performed by transfecting HeLa cells with fluorescently labeled dextran to optimize transfection efficiency; (4) live/dead cell tests were done to assess the viability and health of electroporated HeLa cells at time periods up to 24 hours post treatment. These experiments confirm that NFP electroporation offers single cell selectivity, high transfection efficiency (>95%), qualitative dosage control, and very high viability (92%) of transfected cells. A manuscript in Nano Letters reporting the detailed results, Nanofountain Probe Electroporation of Single Cells, is currently in review. Thus, iNfinitesimal LLC is making great progress toward the development of an instrument for single cell electroporation that is easy to use, precise, and gentle to cells, toward our company’s mission to advance the state-of-the-art in personalized medicine and therapeutics.
