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Updated: Aug 8, 2026
Cell Population Analyses During Skin Carcinogenesis
Published on: August 21, 2013
Quantitative and three-dimensional analysis of human Langerhans cells in epidermal sheets and vertical skin sections
1Department of Clinical Immunology, Karolinska Hospital, Stockholm, Sweden.
Insights
Both epidermal sheets and vertical skin sections are suitable for quantitative analysis of Langerhans cells (LCs). However, epidermal sheets offer clearer 3D reconstructions due to lower background intensity.
Area of Science:
- Immunodermatology
- Cell Biology
- Microscopy
Background:
- Langerhans cells (LCs) are crucial immune cells in the epidermis.
- Quantitative analysis of LCs requires optimal sample preparation methods.
Purpose of the Study:
- To compare the suitability of epidermal sheets versus vertical skin sections for quantitative analysis of Langerhans cells (LCs).
- To evaluate HLA-DR expression and dendritic morphology in LCs using confocal microscopy.
Main Methods:
- Confocal laser scanning microscopy was employed to analyze LCs in normal human skin.
- Samples included acetone-fixed epidermal sheets and 25-micron vertical skin sections.
- Fluorescein isothiocyanate-conjugated anti-HLA-DR antibodies and a threshold algorithm were used.
Main Results:
- No significant differences in epidermal HLA-DR reactivity volume or dendrite number per LC were observed between the two sample types.
- Vertical sections exhibited significantly higher background fluorescence than epidermal sheets.
- Three-dimensional reconstructions showed HLA-DR concentrated in intracellular vesicles in LCs, more frequently in epidermal sheets.
Conclusions:
- Both epidermal sheets and vertical sections are viable for quantitative LC studies.
- Epidermal sheets are preferred for detailed 3D LC analysis due to reduced background noise and larger tissue volume.
- LC dendrite contact was not observed in this study.
Abstract:
We used confocal laser scanning microscopy to analyze and compare Langerhans cells (LCs) in normal skin of six subjects. Acetone-fixed epidermal sheets and 25-microns vertical skin sections were incubated with fluorescein isothiocyanate-conjugated mouse monoclonal antibodies directed against HLA-DR. An individual threshold setting algorithm compensating for the differences in background fluorescence was applied to identify specific fluorescence. No statistically significant difference was found in the relative volume of epidermal HLA-DR reactivity between epidermal sheets (14 +/- 5%) and vertical skin sections (13 +/- 6%) or in the number of dendrites per HLA-DR+ LCs (7.8 +/- 3.1 and 5.9 +/- 3.1, n = 58, respectively). However, statistically significant higher background intensity was found in vertical sections than in epidermal sheets. Three-dimensional (3D) reconstructions of HLA-DR+ LCs revealed a concentration of HLA-DR to one or a few intracellular vesicles in 42 of 58 analyzed LCs in epidermal sheets and in 18 of 58 analyzed LCs in vertical sections. Direct contact between dendrites from different LCs was not found. The results indicate that both skin forms are suitable for quantitative studies. Owing to less background intensity and larger tissue volume, detailed 3D analysis of LCs is preferably performed on epidermal sheets rather than on vertical sections.
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