After After
Before Before

10022 - CD31, Tumor Neovascularization

INTRODUCTION

Angiogenesis, or neovascularization, is the formation of new blood vessels originating from the endothelium of existing vasculature. Angiogenesis is known to play an important role in the neoplastic progression leading to metastasis. CD31 immunohistochemistry (IHC), is widely used in experimental studies quantifying tumor neovascularization in immunocompromised animal models implanted with transformed human cell lines.

By using this APP, neovascularization can be quantified from full virtual slides.

KEYWORDS
Angiogenesis, neovascularization, CD31, microvessel density, cross-sectional area fraction, quantitative, digital pathology, image analysis.

METHODS
All vessel profiles are identified within a region of interest (ROI), which can either be outlined manually (see FIGURE 1) or by an automated method based on image analysis.

Note on area measurements: Only the vessel walls are stained, but the analysis protocol is automatically filling the lumen, in order to get the cross-sectional area measurements for the vessel profiles (see FIGURE 3).

Note on counting: Analysis of full virtual slides takes place in a tile-by-tile fashion. If not handled appropriately, vessel profiles that are intersecting with neighboring tile boundaries would be counted twice (or more). Using an unbiased counting frame, see [1], this can be avoided (see FIGURE 4). This principle is implemented in the present APP. Depending on vessel size and density, the application of this principle could make an important difference.

QUANTITATIVE OUTPUT VARIABLES
Angiogenesis is often quantified as the cross-sectional vessel area fraction and the microvessel density within the region of interest.

The outputs from this protocol are therefore:

- AVES: Total cross-sectional area of CD31-positive vessels

- NVES: Total number of vessel profiles

- AROI: Total area of tissue in ROI

- AA = AVES / AROI: Cross-sectional vessel area fraction

- QA = NVES / AROI: Microvessel density.

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

There are no Auxiliary APPs available.

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. AuthorTM offers a comprehensive and dedicated set of tools for creating new fit-for-purpose analysis APPs, and no programming experience is required.

REFERENCES

LITERATURE
1. Howard, C.V., Reed, M.G. (2005). Unbiased Stereology: Three-dimensional measurement in microscopy. QTP Publications.

RUO
FIGURE 1
FIGURE 1
Outlining the ROI.
FIGURE 2
FIGURE 2
One field of view of the original image at 2X (scaled down to fit this space).
FIGURE 3
FIGURE 3
Classification of the vessels. Note that the lumen of the vessels has been filled in order to get the cross-sectional area measurements for the vessel profiles.
FIGURE 4
FIGURE 4
An un-biased counting frame is included to ensure that each vessel is counted only once.