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10152 - CK16, Skin Damage

INTRODUCTION
Cytokeratin-16 is a type I cytoskeletal protein. It is used as a marker for epidermal hyperplasia, a characteristic which is associated with various inflammatory skin diseases such as atopic dermatitis and psoriasis [1]. In contrast, CK16 protein expression in normal epidermis is very low or absent.

This protocol can be used for quantifying the epidermal expression of CK16. It divides the CK16 staining intensity into four categories: no staining (negative), low staining intensity (1+), medium staining intensity (2+), and high staining intensity (3+). An intensity score is calculated based on the categories.

KEYWORDS
CK16, Skin, Skin damage, Dermatology, Digital pathology, Image analysis, IHC

METHODS
The nuclei present in the epidermis are detected using a polynomial blob filter on a blue-enhancing feature image. The detection allows for exclusion of the nuclei before quantifying the CK16 staining intensity. A feature image, which revolves around red staining, is used for quantification of CK16 intensity. The intensity is divided into four groups ranging from negative (no staining) to strongly positive (3+) based on a threshold classifier and morphological post-processing steps is used for slight smoothing of the stain to handle grainy areas.

QUANTITATIVE OUTPUT VARIABLES
The output variables obtained from this protocol are:

  • Total area of the epidermis excluding nuclei [μm2

  • Percentage of the epidermis area (excluding nuclei) with negative staining intensity

  • Percentage of the epidermis area (excluding nuclei) with 1+ staining intensity

  • Percentage of the epidermis area (excluding nuclei) with 2+ staining intensity

  • Percentage of the epidermis area (excluding nuclei) with 3+ staining intensity

  • Intensity score (inspired by the H score) defined as: 1 x (Percentage of 1+ staining intensity) + 2 x (Percentage of 2+ staining intensity) + 3 x (Percentage of 3+ staining intensity)

AUXILIARY APPs
There are no Auxiliary APPs available.

WORKFLOW
Step 1: Manually outline the epidermis
Step 2: Load and run the APP “10152 – CK16, Skin damage” for the quantification of CK16 staining intensity

STAINING PROTOCOL
There is no staining protocol available.

ADDITIONAL INFORMATION
The APP utilizes the Visiopharm EngineTM and Viewer software modules, where EngineTM offers an execution platform to expand processing capability and speed of image analysis. The Viewer allows a fast review together with several types of image adjustment properties e.g. 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. AuthorTM offers a comprehensive and dedicated set of tools for creating new fit-for-purpose analysis APPs, and no programming experience is required.

REFERENCES
1. De Jong et al., High Expression Levels of Keratinocyte Antimicrobial Proteins in Psoriasis Compared with Atopic Dermatitis, J. Invest Dermatol. 2005, Vol. 125(6), pp. 1163-1173

RUO
Figure 1
Figure 1
Part of epidermis stained with CK16
Figure 2
Figure 2
Manual outline of epidermis
Figure 3
Figure 3
CK16 staining intensity categorization within the epidermis with exclusion of nuclei
Figure 4
Figure 4
Microscopic view of epidermis stained with CK16
Figure 5
Figure 5
Microscopic view of analyzed image