PXD046362

PXD046362 is an original dataset announced via ProteomeXchange.

Title	iCAB: Development of an in situ cell-type specific proteome analysis method using antibody-mediated biotinylation
Description	Since proteins are essential molecules exerting cellular functions, decoding proteome changes is the key to understanding the normal physiology and pathogenesis mechanism of various diseases. However, conventional proteomic studies are often conducted on tissue lumps, in which multiple cell types are entangled, presenting challenges in interpreting the biological dynamics among diverse cell types. While recent cell-specific proteome analysis techniques, like BONCAT, TurboID, and APEX, have emerged, their necessity for genetic modifications limits their usage. The alternative, laser capture microdissection (LCM), although it does not require genetic alterations, is labor-intensive, time-consuming, and requires specialized expertise, making it less suitable for large-scale studies. In this study, we develop the method for in situ cell-type specific proteome analysis using antibody-mediated biotinylation (iCAB), in which we combined immunohistochemistry (IHC) with the biotin-tyramide signal amplification approach. Poly-horseradish peroxidase (HRP) conjugated to the secondary antibody will be localized at a target cell type via a primary antibody specific to the target cell type and biotin-tyramide activated by HRP will biotinylate the nearby proteins. Therefore, the iCAB method can be applied to any tissues that can be used for IHC. As a proof-of-concept, we employed iCAB for mouse brain tissue enriching proteins for neuronal cell bodies, astrocytes, and microglia, followed by identifying the enriched proteins using 16-plex TMT-based proteomics. In total, we identified ~8,400 and ~6,200 proteins from enriched and non-enriched samples, respectively. Most proteins from the enriched samples showed differential expressions when we compared different cell type data, while there were no differentially expressed proteins from non-enriched samples. The cell type enrichment analysis with the increased proteins in respective cell types using Azimuth showed that neuronal cell bodies, astrocytes, and microglia data exhibited Glutamatergic Neuron, Astrocyte and Microglia/Perivascular Macrophage as the representative cell types, respectively. The proteome data of the enriched proteins showed similar subcellular distribution as non-enriched proteins, indicating that the iCAB-proteome is not biased toward any subcellular compartment. Furthermore, we utilized iCAB for cell-type specific proteome analysis in the 5xFAD mouse model. We successfully identified 8164, 8660, 7745, and 7976 proteins from non-enriched sample, neuron, astrocyte, and microglia, respectively. The gene set enrichment analysis enabled us to assess differences in proteome profiling among these cell types, which were not observed in the non-enriched proteome. To our best knowledge, this study represents the first implementation of a cell-type-specific proteome analysis method using an antibody-mediated biotinylation approach. This development paves the way for the routine and widespread use of cell-type-specific proteome analysis. Ultimately, this could accelerate our understanding of biological and pathological phenomena.
HostingRepository	PRIDE
AnnounceDate	2025-05-12
AnnouncementXML	Submission_2025-05-11_21:12:54.429.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	chanhyun na
SpeciesList	scientific name: Mus musculus (Mouse); NCBI TaxID: 10090;
ModificationList	TMT6plex-126 reporter+balance reagent acylated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Fusion Lumos

Revision	Datetime	Status	ChangeLog Entry
0	2023-10-24 10:29:22	ID requested
⏵ 1	2025-05-11 21:12:55	announced

Ryu T, Kim SY, Thuraisamy T, Shin J, Jang Y, Kam TI, Na CH, Overexpression Mouse Model. Adv Sci (Weinh), ():e2409318(2025) [pubmed]

10.1002/advs.202409318;

ProteomeXchange project tag: Human Brain Proteome Project (HUPO_HBPP) (B/D-HPP), Biology/Disease-Driven Human Proteome Project (B/D-HPP), Human Proteome Project

submitter keyword: neuron, astrocytes, microglia., brain, mass spectrometry,Cell-type-specific proteome analysis, immunohistochemistry, biotin-tyramide

Chan-Hyun Na
contact affiliation	associate professor of neurology Institute for cell engineering Johns hopkins university school of medicine
contact email	chanhyun@jhmi.edu
lab head
chanhyun na
contact affiliation	Johns Hopkins University
contact email	chanhyun.na@gmail.com
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2025/05/PXD046362

PRIDE project URI

• PRIDE
  1. PXD046362
     1. Label: PRIDE project
     2. Name: iCAB: Development of an in situ cell-type specific proteome analysis method using antibody-mediated biotinylation