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10020 - Orcein Staining, Skin

INTRODUCTION

Epidermal thickness, and the content of collagen/elastin in the dermal layer, and the relationship to age, gender, skin type, pigmentation, blood content, smoking habits and body site is important in dermatologic research. The epidermal thickness is often affected during skin diseases, such as psoriasis, and its measurement can therefore be a relevant parameter in the assessment of disease progression and treatment effect.
The thickness of the epidermis can vary substantially even within a tissue section. For this reason, the arithmetic mean thickness is estimated as the epidermis area divided by the length of the epidermis. 

KEYWORDS

Skin, Epidermis, Thickness, Psoriasis, quantitative, digital pathology, image analysis, elastin, collagen.

METHODS

Prior to the image analysis, it is important to outline the region of interest, thereby excluding most of the skin, which is not epidermis from the quantitative analysis. This outlining is performed automatically, but could also be done manually. It does not need to be extremely precise, as long as it eliminates tissue areas of the same color as the epidermis. The segmented epidermal layer is skeletonized, thereby generating its midline [See EXAMPLES: figure 3]. The length of the interface between the skeleton and the original epidermis corresponds to two times the length of the epidermis. Finally, a segmentation of the outlined dermis is performed to detect the elastin and collagen content.

QUANTITATIVE OUTPUT VARIABLES

The endpoint for this analysis is the arithmetic mean thickness of the epidermis:

- Epidermal thickness = (Epidermal area) / (Epidermal length)

- Collagen ratio = (Collagen area) / (Dermis area)

- Elastin ratio = (Elastin area) / (Dermis area)

The analysis could be extended to also include area ratio (epidermis to total tissue ratio) or other relevant measures.

AUXILIARY APPS (included)

Auxiliary APPs are used for additional process steps, e.g. finding Region of Interest (ROI).

TISSUE DETECT

The auxiliary APP '01 Tissue ROI' can be used to automatically detect the region of interest (ROI). This is to save time in the subsequent analysis steps. The analysis is carried out at a lower magnification and finds larger pieces of tissue. Dust and small artifacts are ignored. There are no quantitative output variables from this step [See EXAMPLES: figure 1 and 2].

ELASTIN/COLLAGEN QUANTIFICATION

The auxiliary APP '03 Analyze Collagen Elastin' segments the detected dermis into different components: collagen and elastin. 


STAINING PROTOCOL

There is no staining protocol available

ADDITIONAL INFORMATION

The APP utilizes the EngineTM and Viewer software modules, where EngineTM offers an execution platform to expand processing capability and speed of image analysis. Viewer gives a fast review together with several types of image adjustment properties ex. outlining of regions, annotations and direct measures of distance, curve length, radius, etc. 
By adding the AuthorTM module the APP can be customized to fit other purposes. AuthorTMoffers a comprehensive and dedicated set of tools for creating new fit-for-purpose analysis APPs, and no programming experience is required.

Workflow

Step 1: Load auxiliary APP for tissue detection '01 Tissue ROI' and run on the image to process. This outlines the tissue with a Region Of Interest (ROI) and saves time in the subsequent analysis steps.

Step 2: Load quantification APP '02 Analyze' and run on the image to detect the epidermis and calculate its thickness.

Step 3 (optional): Load quantification APP '03 Analyze Collagen Elastin' and run on the images to segment the dermis into collagen and elastin.

REFERENCES

There are currently no references available.

RUO
FIGURE 1
FIGURE 1
Orcein stained skin section.
FIGURE 2
FIGURE 2
Orcein stained skin section, with rough outlining of the epidermis. This is performed automatically.
FIGURE 3
FIGURE 3
The area (red) and length (green) of the epidermis is used for calculating its arithmetic mean thickness.
FIGURE 4
FIGURE 4
Additional detection of the elastin (blue) and collagen (yellow) in the skin section. This is only performed in the dermis.