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D E F I N I T I O N S

The definitions of different parameters of the comet are dependent on the software one is using:

 

For KometTM software the definitions are as follows


A typical comet showing analysis cell with head, tail and Background ROIs
(ROIs - regions of interest)

Tail Length: Tail Length is the distance of DNA migration from the body of the nuclear core and it is used to evaluate the extent of DNa damage.
Tail Length = Tail Extent
(Tail from Center) Tail Extent + Head Extent/2

Olive Tail Moment: Tail moment is defined as the product of the tail length and the fraction of total DNA in the tail. Tail moment incorporates a measure of both the smallest detectable size of migrating DNA (reflected in the comet tail length) and the number of relaxed / broken pieces (represented by the intensity of DNA in the tail).

Olive Tail Moment=(Tail.mean - Head.mean) X Tail%DNA/100.

Extent Tail Moment=Tail Length X Tail%DNA/100.

Head % DNA = (Head.OptInten /(Head.OptInten + Tail.OptInten)) X 100.

Tail % DNA = 100 - Head%DNA.

 

For Vis CometTM software the definitions are as follows:

1          Comet extent
2          Comet total intensity
3          Comet total area
4          Tail extent
5          Tail extent moment
6          Tail (Olive) moment
7          Tail integrated intensity (Singh)
8          Tail integrated intensity ratio (Singh)
9          Tail area (Singh)
10        Tail break Number (Singh)
                

1) Comet Extent
The extent is the distance from the leading edge to the trailing edge and defined as follows.

2) Comet total intensity
Sums all pixel intensities from the grayscale image when there is a corresponding pixel in the binary mask image. The resulting value is normalized by the maximum intensity value that can be represented in the data buffer.
Example:
When working with 8-bit images, the maximum intensity value can be 255. Working with 12-bit images, the maximum intensity can be 4095. In case of an 8-bit image, the total intensity would be:

3) Comet total area
The number of pixels in the binary comet image (including pixels that belong to fragments) is counted. The area is returned in calibrated units.

5) Tail extent moment
It is defined as follows:  

  L= length of the tail or body
DNA = DNA of tail or body given as a percentage

6) Olive Moment
It is defined as follows:

 
CG = center of gravity of the tail or body weighted by gray values
CGH = center of gravity of the head weighted by gray values
DNA = tail or body DNA

7) Integrated intensity
The integrated intensity is a feature, that incorporates different geometric parameters. Due to the fact, that damaged cells change their length and breadth, exactly these two geometric parameters are used for the calculation.

The algorithm starts from the beginning of the tail and examines each vertical scan line until the end of the tail is reached. Each vertical scan line is processed from top to bottom and only those pixels are considered that are identified as signal pixels.

From each vertical scan line the product of breadth, position and total intensity is accumulated.

The integrated intensity is calculated as follows:

 
Ip is the product of breadth, position and total intensity of one column

8.) Tail Intensity Ratio:
The integrated intensity ratio is calculated as follows:


where
IT = sum of all intensity values
The figure below demonstrates the mechanism for the calculation.

 


9)Tail Area:

The number of pixels in the binary tail region of image (including pixels that belong to fragments) is counted. The area is returned in calibrated units.

10) Tail Breaks Number:
All objects (or sometimes called blobs or fragments) in the tail area (red overlay area) are counted. This number minus 1 is number of  DNA breaks.

 

Courtesy: Dr. N.P. Singh