Immuno Tomography (IT) and Imaging Mass Cytometry (IMC) for constructing spatially resolved, multiplexed 3D IMC data...

Area of Science:

• Biomedical Engineering
• Stem Cell Biology
• Ophthalmology

Background:

• Immuno Tomography (IT) identifies label-retaining stem cells in cornea and meibomian glands.
• IT quantifies small stem cell populations (1-4 cells) but is limited by antigen stability for multi-antigen characterization.

Purpose of the Study:

• Evaluate Imaging Mass Cytometry (IMC) for multiplexed imaging of Immuno Tomography (IT) plastic sections.
• Generate 3-dimensional IMC (3D IMC) data sets for enhanced stem cell analysis.

Main Methods:

• Utilized K5-H2B-GFP mice eyelid tissue processed for IT.
• Probed 400 serial plastic sections (2 μm) with 8 metal-tagged antibodies (sox 9, collagen type I, E-cadherin, Ki67, GFP, αSMA, vimentin, DNA intercalator).
• Generated multiplexed images using an IMC system and assembled 3D reconstructions.

Main Results:

• Successfully detected all 8 metal-labeled tags and assembled images into 3D IMC datasets.
• Identified GFP-labeled nuclei in meibomian glands, consistent with previously reported slow-cycling stem cell numbers.

Conclusions:

• IMC effectively generates multiplexed, 3D data sets from plastic sections for stem cell localization.
• 3D IMC shows potential for comprehensive characterization of stem cell populations across various tissues.