Slide Scanning

Basis for virtual microscopy are virtual slides (also called “WSI-Whole Slide Images”). They are generated through high-resolution scanning of glass slides, done by slide scanners. Such devices are available from several vendors, but the basic principle is roughly the same:

  • The glass slide is mounted into a frame
  • The frame is positioned under an objective lens
  • By means of a digital camera, the first tile is focussed automatically 
  • The camera saves the image 
  • The frame is positioned to the next field for saving the next image tile
  • This way hundreds to thousands of images are generated, which together represent the whole tissue probe in highest magnification 
  • All image tiles are stitched together to one image 
  • Finally the image is saved in a suitable, mostly compressed image format 

Virtual Slides

Virtual Slides have image dimensions of up to 200,000 times 100,000 pixels. Uncompressed, they are up to 40 GB in file size, compressed up to 2 GB. Biopsies would need about 400 MB on average, TMAs 1 GB and ectomies about 1.6 GB. On average for all slides you can calculate with 1 GB per slide. 

Virtual slides are stored in different formats, such as JPEG2000 (do not mix with JPEG) or in vendor specific formats. Often a virtual slide consists of a whole folder with several files, like in our “VSF” format. The reason for that is to support a faster access on single image tiles.magnification

Resolution

Most slide scanners generate images with a pixel resolution of about 0.3 Ám in x and y-direction. That means, a pixel will cover an area of about 0.3 Ám times 0.3 Ám in nature (on the tissue probe).

Viewed on a monitor (with a resolution of about 0.23 Ám x 0.23 Ám per pixel) that leads to a calculative magnification of about 650-fold. But because the resolution of a monitor is limited, the image does not appear as clear as in the light microscope. What you really can interpret roughly corresponds to 400-fold magnification, which means 40 x objective lens.

The resolution of a slide is not only defined through the objective lens during scanning. The CCD detector of the camera mostly has such a high density, that it also adds to the resolution. As result, “20” x objective in a slide scanner does not mean, the resulting images would only be 200-fold magnification.

Image Commmunication

Virtual slides have a file size of about 1 GB compressed on average. With that file size, a fast and effective transmission of the whole image over internet is impossible. Even on a computer, such an image could not be loaded to the RAM memory completely.

It needs certain methods to access virtual slides. That is the so called “Image Streaming”. The basic idea is to leave the original image on the server, and transmit only that image region, which is currently needed on the client side. The server extracts only the current region in the current magnification from the original image, compresses and transmits to the client, which shows it in the virtual microscope. That reduces the data communication to small image tiles, which is comparable to the network traffic of general internet browsing.

Visualisation

The visualisation of virtual slides by the user is done using the virtual microscope. That application at least simulates the functionality of a light microscope (changing of the objective, moving, selecting regions). Furthermore, it mostly offers several functions like measuring, annotations, saving and many more.

 

Screenshot of VM Slide Explorer

VM Slide Explorer Software

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