Molecularly Defined Immunoprobes for Scalable Brain Protein Mapping with Reference Datasets

University Of California At Davis, Davis CA

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Abstract

ABSTRACT Molecularly Defined Immunoprobes for Scalable Brain Protein Mapping with Reference Datasets To fully understand brain function and dysfunction, it is essential to map the brain's circuitry from molecules to cells to circuits. Recent advancements in whole brain tissue clearing have opened new avenues for cellular mapping at the proteomic level. However, the lack of validated, molecularly defined, renewable reagents has limited progress in this area. Monoclonal antibodies (mAbs) serve as effective affinity probes for scalable whole brain proteomic mapping. Developing a renewable resource of monospecific immunolabelers specifically validated for whole brain cellular mapping represents a significant step forward in creating reference atlases of the brain. Our project aims to enhance the dissemination of an existing monoclonal antibody collection by identifying mAbs that effectively label target proteins in both mouse and human cleared tissues. We will convert these mAbs into recombinant forms (R-mAbs) for use in generating comprehensive reference datasets for whole mouse brains. Current limitations in cleared brain immunolabeling hinder the utility of existing antibody collections, which have not been optimized for these new methodologies. The extensive repository of cryopreserved hybridomas at the UC Davis/NeuroMab facility, targeting over 500 brain proteins, provides a unique opportunity to broaden the applicability of this collection to cleared whole brain mapping. Collaborating with Dr. Zhuhao Wu at Cornell, an expert in whole brain protein mapping, we will identify and validate probes that bind to targets in both mouse and human cleared brain tissue. Our approach includes validating sets of R-mAbs for cleared tissue labeling and, in collaboration with AddGene, disseminating these plasmids and R-mAbs widely within the scientific community. Furthermore, we will generate a reference dataset for at least two R-mAbs per target in cleared whole mouse brain, which will be made accessible through a cloud-based platform. This initiative will significantly enhance the availability of these critical resources and accelerate the pace of brain circuit mapping at various scales.