Scalable spatial single-cell transcriptomics and translatomics in 3D thick tissue blocks

Area of Science:

• Molecular Biology
• Genomics
• Biotechnology

Background:

• Understanding gene expression in 3D tissue context is crucial for biology and disease.
• Existing spatial profiling methods are limited to thin tissue sections (5-20 µm).

Purpose of the Study:

• To develop novel 3D in situ spatial profiling techniques for gene and translation activity.
• To enable high-resolution analysis of gene expression within thicker tissue blocks.

Main Methods:

• Developed Deep-STARmap for transcript quantification and Deep-RIBOmap for translation activity.
• Utilized scalable probe synthesis, hydrogel embedding, and cDNA crosslinking.
• Integrated multicolor fluorescent protein imaging for cell typing and morphology tracing.

Main Results:

• Achieved 3D in situ quantification of thousands of gene transcripts and translation activities in 60-200 µm thick tissues.
• Successfully performed molecular cell typing and 3D neuron morphology tracing in mouse brain.
• Demonstrated comprehensive analysis of tumor-immune interactions in human skin cancer.

Conclusions:

• Deep-STARmap and Deep-RIBOmap overcome limitations of existing methods, enabling deeper tissue analysis.
• These techniques provide unprecedented insights into gene function in complex 3D tissue environments.
• 3D spatial profiling is valuable for studying tissue structure, function, and disease pathology.