This Small Business Innovation Research (SBIR) Phase I project proposes the development of a new imaging platform based on mass spectrometry that addresses the need for a technology with high spatial resolution and sensitivity in the low-mass range. Mass spectrometry imaging (MSI) is a rapidly growing area because it does not require a labeled sample, offers location-specific chemical information, and has high information content with potentially thousands of compounds detected at each pixel in the image. However, broad-based applicability of MSI has not been realized due to limited spatial resolution and sensitivity in the low-mass range. The overall Phase I goal is to overcome the current MSI limitations and expand the applicability by utilizing nanostructured surfaces that take advantage of an entirely different desorption/ionization mechanism than current surface-based mass analysis technologies. Our Phase I objective is to develop two prototypes based on this nanostructured surface. First, we will develop a prototype for mass-based imaging with high spatial resolution and high low-mass sensitivity that can be used for tissue imaging. Second, we will produce a prototype that enables a new approach to high-throughput screening of enzymatic activity libraries based on mass imaging, with a projected throughput of more than 10,000 assays/day.
The broader impact/commercial potential of this project is that mass-based imaging can be used in applications that are difficult or impossible with the current MSI technologies, which includes the matrix-assisted laser desorption/ionization (MALDI) and secondary-ion mass spectrometry (SIMS) technologies. The ability to perform MSI in the low-mass range will have particularly high impact because many important classes of molecules fall within this range including biofuels, therapeutic compounds, disease-relevant metabolites, environmental toxins, food contaminants, and industrial compounds. Societal and commercially important mass-based imaging applications of the proposed platform include 1) animal/plant tissue imaging for drug discovery, drug localization/metabolism, disease marker discovery, environmental studies and 2) biofilm imaging for biofuel production and environmental remediation. The proposed MSI-based high-throughput screening platform has significant potential for discovery of new enzymes for biofuel production industrial processes as well as therapeutic enzyme inhibitors. In addition, the nanostructured surfaces used in this proposal have a low barrier of entry and high value to the customer because the end user can take advantage of the new capabilities enabled by the proposed platform, while using the same commercially available laser desorption/ionization mass spectrometer that they currently use for matrix-assisted laser/desorption ionization (MALDI).
The two key aims of this SBIR Phase I project were: 1) Develop a mass spectrometry imaging (MSI) surface with 15 micron or better spatial resolution and high sensitivity for metabolites. 2) Develop a mass spectrometry imaging (MSI) based high-throughput screening platform with the capability to analyze at least 10,000 samples per day. To achieve aim 1, a new enhanced-activity MSI surface was developed to enable high sensitivity detection of metabolites at an image resolution of 5 microns. In order to demonstrate the successful achievement of Aim 1, single cells were spotted on the enhanced MSI surface and the spots were imaged at 5 micron resolution. At this resolution, high-quality metabolite-focused mass spectra from single isolated cells on the surface were obtained allowing us to begin to observed metabolic heterogeneity between single cells, which is difficult or impossible with traditional MSI and mass spectrometry technologies. This particular application is highly relevant in the pharmaceutical/diagnostic markets as well as the industrial biological production markets. To achieve aim 2, we developed a new integration method for coupling picoliter volume acoustic array deposition to mass spectrometry imaging (Refer to Attached Image). As a result of this integration, in addition to further enhancements of our MSI surface, we have developed a platform capable of high-throughput analysis of more than 20,000 samples/day while requiring only picoliter volumes of sample and generating almost zero waste. In order to demonstrate successful achievement of Aim 2, 18,000 cellulase activity assay samples were printed and analyzed using this new MSI platform at throughput rates that exceed 20,000 samples per day. This demonstration is particularly relevant given that the discovery of new cellulase activity is critical for efficient biofuel production. The broader impact of this work is to develop a technology that enables the high-throughput analysis and discovery of new chemical and biological reactions that are currently inaccessible or not well served with existing analytical technologies. This includes the discovery of new enzymes and active molecules for biofuel, industrial, environmental, pharmaceutical, diagnostic, and quality control applications. In addition, the high-throughput nature of this MSI technology reduces overall R&D costs for chemical/biological discovery/analysis and reduces the hazardous waste generated during the analysis to nearly zero. As a result of successful achievement of Phase I aims, we have formed a new company, Nextval Inc. that is focused on the commercialization of high-throughput mass spectrometry imaging technologies and applications. Nextval has already landed several commercial contracts based on technologies derived from this SBIR Phase I project.
