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Updated: May 27, 2026
In Vivo Two-Color 2-Photon Imaging of Genetically-Tagged Reporter Cells in the Skin
Published on: July 11, 2019
Real-time visualization of macromolecule uptake by epidermal Langerhans cells in living animals
Rachel E Frugé1, Colleen Krout, Ran Lu
1Department of Medical Microbiology and Immunology, University of Toledo College of Medicine, Toledo, Ohio 43614-5806, USA.
Insights
Researchers developed a new imaging method to visualize Langerhans cell (LC) endocytosis in living animals. This technique tracks how LCs internalize substances, aiding in the study of skin immunity and antigen presentation.
Area of Science:
- Immunology
- Cell Biology
- Dermatology
Background:
- Langerhans cells (LCs) are key skin-resident immune cells involved in antigen presentation.
- Understanding LC endocytotic activity is crucial for studying skin immunity.
Purpose of the Study:
- To develop and validate a novel in vivo imaging method for assessing Langerhans cell endocytosis.
- To visualize and characterize the endocytotic processes of LCs in living mice.
Main Methods:
- Subcutaneous injection of FITC-conjugated dextran probes into mouse ear skin.
- Confocal microscopy to detect and quantify fluorescently labeled epidermal cells.
- Intracellular localization studies using Alexa Fluor 546-conjugated probes in genetically modified mice (IAβ-EGFP and Langerin-EGFP-DTR).
Main Results:
- FITC-positive epidermal cells, identified as LCs (>90%), were detectable 12-24 hours post-injection.
- Internalized dextran probes showed close association with MHC class II molecules but not with Langerin.
- The imaging approach successfully monitored LC function in wild-type animals.
Conclusions:
- The developed imaging method effectively visualizes Langerhans cell endocytosis in living animals.
- LCs continuously sample their environment, internalizing various materials.
- This technique offers a valuable tool for studying LC function and skin immune responses.
Abstract:
As a skin-resident member of the dendritic cell family, Langerhans cells (LCs) are generally regarded to function as professional antigen-presenting cells. Here we report a simple method to visualize the endocytotic activity of LCs in living animals. BALB/c mice received subcutaneous injection of FITC-conjugated dextran (DX) probes into the ear skin and were then examined under confocal microscopy. Large numbers of FITC(+) epidermal cells became detectable 12-24 hours after injection as background fluorescence signals began to disappear. Most (>90%) of the FITC(+) epidermal cells expressed Langerin, and >95% of Langerin(+) epidermal cells exhibited significant FITC signals. To assess intracellular localization, Alexa Fluor 546-conjugated DX probes were locally injected into IAβ-enhanced green fluorescent protein (EGFP) knock-in mice and Langerin-EGFP-diphtheria toxin receptor mice--three dimensional rotation images showed close association of most of the internalized DX probes with major histocompatibility complex (MHC) class II molecules, but not with Langerin molecules. These observations support the current view that LCs constantly sample surrounding materials, including harmful and innocuous antigens, at the environmental interface. Our data also validate the potential utility of the newly developed imaging approach to monitor LC function in wild-type animals.
