Abstract Tremendous progress has been made in defining Alzheimer’s disease (AD)-associated cell types by single-cell sequencing approaches, such as scRNA-seq. However, the nucleic-acid–based methods are blind to proteins. Therefore, AD’s proteome and post-translational modifications (PTMs) remain largely unknown. Another limitation of single-cell sequencing is that the standard cell isolation techniques damage the fine projections of brain cells, such as the dendrites of neurons. Missing biomolecules in these brain-cell-specific structures will seriously mislead our understanding of molecular mechanisms of AD progression. There is an urgent need for new technologies to fill the technical gaps in the proteomics and multiomics analysis of AD. Here, we bring to the AD field our NIH Director’s New Investigator Award-winning method, Gel-based Optical-isolation 3D (GO3D) multiomics. It can precisely dissect tiny projections of brain cells from whole-mount brains for subsequent mass spectrometry analysis and next-generation sequencing. This method is an ideal approach to achieving my long- term goal: creating a single-cell-type multiomics map of AD. As the first step, this project focuses on one critical AD-associated cell type, the astrocyte. We will use GO3D single-cell-type multiomics to identify AD-associated proteins and PTMs in astrocytes (Aim 1) and establish a multiomics map of AD featuring astrocytes (Aim 2). 5xFAD mouse models at different time points will be used to examine AD in the early, middle, and late stages. Human prefrontal cortex tissues will be used to validate key discoveries made from mice. Unbiased profiling of the proteins together with RNAs and chromatin accessibility in astrocytes will provide a list of AD-associated proteins and PTMs and their regulatory mechanisms, and thus will greatly advance our understanding of the molecular basis of AD.