Microscope image processing
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Microscope image processing is a broad term that covers the use of digital image processing techniques to process, analyze and present images obtained from a microscope. Such processing is now commonplace in a number of diverse fields such as medicine, biological research, cancer research, drug testing, metallurgy, etc. A number of manufacturers of microscopes now specifically design in features that allow interface to an image processing system.
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Image acquisition
Acquisition is usually done using a CCD camera mounted in the optical path of the microscope. The camera may be full colour or monochrome. Very often, very high resolution cameras are employed to gain as much direct information as possible. Cryogenic cooling is also common, to minimise noise. Often digital cameras used for this application provide pixel intensity data to a resolution of 12-16 bits, much higher than is used in consumer imaging products.
In recent years, much effort has been put into acquiring data at video rates, or higher (30 frames per second or higher). This allows dynamic processes to be observed in real time, or stored for later playback and analysis. Combined with the high image resolution, this approach can generate vast quantities of raw data, which can be a challenge to deal with, even with a modern computer system.
2D Image techniques
Techniques employed for processing microscopy images are in general rather different from those encountered in consumer image processing (e.g. Photoshop) type applications, though most applications will have a suite of standard operations such as sharpening, contrast and brightness enhancement, and colour balancing. One key requirement is to electronically "undo" the distortion of the optical path of the microscope, thus eliminating distortions and blurring caused by the instrumentation. This process is called deconvolution, and a variety of algorithms have been developed, some of great mathematical complexity. The end result is an image far sharper and clearer than could be obtained in the optical domain alone. A typical software package for 2D microscopy is Improvision's Openlab, others include NIH Image, LabVIEW, etc.
3D image techniques
Another common requirement is to take a series of images at a fixed position, but at different focal depths. Since most microscopic samples are essentially transparent, and the depth of field of the focused sample is exceptionally narrow, it is possible to capture images "through" a three-dimensional object using 2D equipment. Software is then able to resconstruct a 3D model of the original sample which may be manipulated appropriately. The processing turns a 2D instrument into a 3D instrument, which would not otherwise exist. In recent times this technique has led to a number of scientific discoveries in cell biology. Some software packages for 3D microscopy are Improvision's Volocity and Scientific Volume Imaging's Huygens Software.
Analysis
Analysis of images will vary considerably according to application. Typical analysis includes determining where the edges of an object are, counting similar objects, calculating the area, perimeter length and other useful measurements of each object. A common approach is to create an image mask which only includes pixels that match certain criteria, then perform simpler scanning operations on the resulting mask. It is also possible to label objects and track their motion over a series of frames in a video sequence.
External Link
- Confocal Microscopy: Troubleshooting Image Quality (http://www.confocal-microscopy.org/Troubleshooting.htm)