SBIR-STTR Award

This Small Business Innovation Research (SBIR) Phase I project proposes to construct a new microarray platform with high protein binding capacity that allows for enhanced fluorescence detection. Limitations of existing microarray surfaces include platform-based optical interferences and limited or ineffective binding capacity for biomolecules. These limit the ultimate sensitivity of binding reactions mounted on existing microarray surfaces. We will construct a composite or modified surface in two ways. First, by casting nitrocellulose (a polymer able to bind many different biomolecules essentially irreversibly) on an optically transparent porous track-etched membrane. We believe this new surface will maintain some properties of both starting materials. By varying the pore structure of the track-etched membrane, we will optimize the resulting membranes ability to capture complex protein mixtures and permit sensitive fluorescent detection. Secondly, we will directly modify the track-etched membrane with functional silanes to provide chemical groups permitting covalent coupling of proteins and nucleic acids. This approach should prove beneficial to maintain the optical compatibility of the original track-etched structure. This type of modified, optically transparent track-etched membrane may be optimal for antibody arrays where the capture molecule can be immobilized at a sufficient density to provide a sensitive assay surface. The broader impact/commercial potential of this project will be to provide the basis for a new analytical tool that will allow establishment of antibody- and antigen-based assays of enhanced sensitivity. It also will provide an innovative surface to capture quantitatively the biochemical components of complex mixtures in such a way as to permit the detection of rare molecules. Microarrays play an increasingly important role in bioscience research, disease, and drug discovery processes as well as in human and animal diagnostics. They provide parallel processing tools required to extract multiple values from small amounts of precious clinical and research samples. Microarrays with enhanced sensitivity will permit the detection of rare biomolecules that may be involved in cellular regulation, cellular differentiation, and disease mechanisms. Reverse phase protein arrays (RPPA) are important for understanding cellular changes in a variety of disease states. In cancer, for example, lysates from small numbers of tumor cells can be spotted on a surface and then interrogated with many different antibodies to elucidate protein expression patterns in these tumor cell populations. The power of these techniques will be enhanced significantly by having a platform able to support the most sensitive assays.

This Small Business Innovation Research (SBIR) Phase II project will provide an optimized composite polymer protein binding surface for proteomics applications. The new surface will be specifically designed for reverse phase protein microarrays to enable detection of rare molecules in complex biological mixtures. Discovery and quantification of rare molecules in complex mixtures is essential to improve the understanding of disease mechanism and progression, and responses to treatment regimes. Current surfaces used in these applications have properties that exhibit limited sensitivity of detection due to optical interferences, low protein binding and accumulation of nonspecific interactions. This project will optimize and introduce the application of a new track etched , nitrocellulose composite membrane for protein array applications. Manufacturing processes for the new composite will be developed to generate multiple forms of the composite to allow it to be incorporated into a variety of binding assay formats. This effort will also shed light on important properties for generating ultrasensitive binding surfaces. The result of this project will be an optimized composite membrane with characteristics and manufacturability suited for the most sensitive binding applications, such as reverse phase protein arrays. The platform initially will be optimized for fluorescent detection of rare molecules in complex cell lysates. The broader impact/commercial potential of this project will be to provide a family of discovery and diagnostic tools that will expand the understanding, detection and treatment of human disease. The current focus in translational medicine for therapies in clinical trials is to identify expression patterns of proteins (biomarkers) in individual patients. These measurements allow the monitoring and understanding of individualized disease progression and responses to treatment. They will provide the data necessary to create targeted, personalized treatment regimens. Protein arrays have found utility over the past decade as research tools that provide multiplexed detection and quantification of protein expression. However, the full potential of these tools as diagnostic platforms that provide patient-specific information and guide drug treatment has not been realized due to insufficient binding capacity, limited dynamic range and poor sensitivity. This project defines a new composite surface that has a significant increase in both binding capacity and sensitivity when incorporated into multiplexed immunoassay systems. The composite can be included in a variety of platforms to enhance discovery and quantification of important markers on an individual scale as well as high throughput systems for broad diagnostic application