The objective of this proposal is to attempt a new mode of analysis for biological tissue by matrix-assisted laser desorption ionization imaging mass spectrometry (MALDI-IMS). MALDI-IMS is a powerful analytical technique capable of generating ~ 50-100 micrometer resolution images that can be filtered by mass, directly from biological tissue sections. Applications for the technique range from cataloging the distribution of bio-molecules and pharmaceuticals in tissues, to biomarker discovery, tumor classification, elucidation of pathogenesis, studies of developmental biology, etc. This proposal outlines a new technology aimed at improving MALDI-IMS analyses on several fronts. The basis of the technology is the construction of novel substrates designed to selectively extract, capture and enrich analyte(s) of interest from tissue, while preserving sample spatial resolution, in a format that will interface directly with the mass spectrometer. The methodology outlined here-in directly addresses the most recognized and consistent points of weakness in current MALDI-IMS analyses and will serve to enhance MALDI-IMS performance by providing more detail, balance and versatility to existing applications as well as providing exciting potential for expansion.
The specific aims for this initial phase of the project target substrate construction and experimental procedure.
Each aim i s critical to proof- of-product feasibility and they allow for quantifiable evaluation and comparative performance with current, state-of-the-art MALDI-IMS technology. It is hypothesized here that the proposed technology will increase the detection thresholds of MALDI-IMS by enabling selectivity for targeted analyte(s) with simultaneous removal of interferents. It will increase sensitivity by allowing more appropriate conditions for analyte / matrix co-crystallization and it will increase data quality by providing an environment more optimal for mass spectrometry. It will stabilize the tissue image by presenting a substrate that will accurately lock-in the spatial integrity of the tissue at a resolution equal-to or greater- thn what is achievable by current protocols. It will increase intra-sample consistency and inter-sample reproducibility by decoupling the steps of analyst extraction and matrix deposition and providing a more systematic and optimized means for accomplishing these tasks. It will expand experimental capability by allowing sample manipulation without disturbing the spatial integrity. It will expand experimental capability by preserving the parent sample allowing for multiple, repetitive or divergent analyses to be performed on the same specimen. It will expand experimental capability by enabling the analysis of tissues of diverse shapes and forms and even allow for the analysis of living tissue. It will simplify experimental protocol by removing restrictions for cryogenically sliced tissue specimens and it will hopefully serve to expand participation in MALDI-IMS technology by lifting the need for specialized and expensive equipment. The prototype substrates are composed of a Duocel(R) reticulated vitreous carbon (RVC) foam combined with a porous polymer monolith. The materials used to develop these unique substrates are commercially available. The chemistry to be employed in plate construction is well understood and the reactants needed are commercially available. The tissue sample manipulations that will be attempted are documented in the literature. The preliminary data demonstrated here-in are encouraging.
This proposal outlines a new technology aimed at improving the quality and expanding the capability of tissue imaging by matrix-assisted laser desorption ionization mass spectrometry (MALDI- IMS). MALDI-IMS is already a powerful tool, widely-used in biological and biomedical research, for generating high spatial-resolution, molecular-level images of biological tissues. The application of MALDI-IMS has significantly and positively impacted research and understanding of menacing diseases and wondrous events in developmental biology. This proposal describes an innovative mode for performing MALDI-IMS that aims to allow more detail, precision and versatility in existing applications, and offer potential for expansion in the utility and number of practitioners of the technique.