Spatial Proteomics toward Subcellular Resolution by Coupling Deep Ultraviolet Laser Ablation with Nanodroplet Sample...

Area of Science:

• Proteomics
• Molecular Biology
• Biotechnology

Background:

• Spatial omics provides critical insights into cellular functions and disease pathology.
• Current laser microdissection methods for spatial proteomics are limited by low throughput and resolution.
• Subcellular resolution (<10 μm) is needed for advanced proteomics studies.

Purpose of the Study:

• To develop a high-throughput and high-resolution spatial proteomics platform.
• To overcome the limitations of existing laser microdissection techniques.
• To enable detailed molecular profiling of subcellular regions in tissue samples.

Main Methods:

• Coupling of deep ultraviolet (DUV) laser ablation (LA) with nanoPOTS (Nanodroplet Processing in One pot for Trace Samples)-based sample preparation.
• Development of a proximity aerosol collection approach for improved sample recovery.
• Analysis of human pancreas tissue sections with varying ablation spot sizes.

Main Results:

• The DUV-LA system achieved a throughput of ~30 spots/min and a spatial resolution down to 2 μm.
• The DUV-LA-nanoPOTS platform detected an average of 1312, 1533, and 1966 proteins from spots of 7, 13, and 19 μm diameters, respectively.
• Proteins specifically enriched to subcellular compartments were identified in a proof-of-concept study.

Conclusions:

• The developed DUV-LA-nanoPOTS platform offers a high-throughput and high-resolution solution for spatial proteomics.
• This platform enables deep proteomic mapping at the subcellular level.
• It facilitates the study of cellular heterogeneity and disease mechanisms through detailed spatial molecular profiling.