Help! There is a comet in my computer! 38
5.7 Why a 10-bit camera beats an 8-bit camera
In the comet assay, a camera serves as a detector of light intensity and hence its properties
affect the quality of the measurement. There are many different types of cameras out there,
and we will only cover some topics essential for understanding the comet measurement.
We have video cameras and digital cameras, and both can be used for the comet assay.
They differ by the mode of data transfer from the camera to the computer, but this is of no
concern here. Then, we have colour and monochrome (“black and white”) cameras. A
monochrome camera is a better tool for detection of fluorescence – anyway, we are only
measuring light intensity, not wavelength (our signal is a narrow band of orange to red
wavelengths). The fluorescence signal is very weak, hence the camera has to have a high
sensitivity – it has to be sensitive to low light intensities. It is also better if a camera has a
low noise. Noise is a signal, generated by various components of the camera, which is not
related to the true signal from the slide. A camera used for the comet assay must have a
linear response to light intensity, according to our postulate in Figure 4. Some cameras
show considerable deviation from linearity and are not suitable for the comet assay.
Cameras also have different resolutions – the number of light-sensitive elements in the
camera detector.
Finally, cameras differ in the way they record pixel grey values. So far we always assumed
that the camera records 256 different grey values. However, this is only true for an 8-bit
camera. Pixels can also be described with a higher number of grey values: 1024 grey
values in 10-bit cameras, 4096 in 12-bit cameras, 16384 in 14-bit cameras and so on. The
number of available grey values to describe each pixel is called pixel depth.
If your camera has pixel depth higher than 8-bit, then all of the examples described in
previous chapters also hold true for your camera, except that wherever we used 256 grey
values you have to imagine a higher number of grey values.
Since our eyes detect much fewer shades of grey than 256, we do not need to use pixel
depth higher than 8-bit if we are only recording images to look at. However, there are some
advantages in using cameras with a higher pixel depth when we are recording images for
measurements of light intensity.
Let us compare an 8-bit and a 10-bit camera used for the comet assay. We have either 256
or 1024 available grey values. Let us record an image with an 8-bit camera (Figure 23A).
In this image, a large part of the available grey values is used to record the signal from the
slide. A 10-bit image has four times more available grey values, so let us scale up the x-
axis in the histogram of image grey values to 1024 (Figure 23B). We see that if we record
a 10-bit image that contains the same amount of information as our original 8-bit image, a
very small part of the 10-bit range of available grey values is used (Figure 23C). So when
we record a good quality image with a 10-bit camera, we have more grey values available
to record the signal, and we can record light intensity with a higher precision (Figure 23D).
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