Dear sir,

try to use CASP. It's free software, which gives you the opportunity to get quite accurate results. Yoy can find it on website http://casplab.com/download.

Best regards,

Maxim

Image J is a very versatile software package that can be used to analyse everything from microscopic images to DNA dot-blots. I have used it very successfully to do FRAP in homogenous gels. For the Comet Assay, you can define Regions of Interest (the head and the tail), measure lengths, areas and grayscale for all ROIs, and subtract background. This can be used to calculate parameters such as Percentage of DNA in the Tail and Tail Extent Moment.

Dear Sally,

I think "Image J" isn't usefull software for the DNA-comet assay. This software counts parameters no automatically, and it can be confusing for some researchers. So CASP counts all comet's parameters in "one click".

Dear Maxim and Sally,

I appreciate your kind responses.

Reading your answer and Sally's, I was thinking: can I use CASP to analyze some parameters in comet assay and complement with Image J?

I searched in CASP and couldn't find any feature of the software to calculate % of DNA in the tail.

Is that possible to assess this parameters stained with silver nitrate? I saw only in a fluorescent stain...

Best regards,

Giuseppe.

Hello,

might you attach your comet's image?

This is a test from last week.

I've tried to analyze your image by CASP, but I couldn't do it. The software can't identify the comet head. What about modification of the silver staining protocol? I attached the article with some suggestions about this procedure.

I have performed the comet assay, but used SyBr green for detecting tail moment. A free software, Comet Score, is available online. I don't know if this would also work for your silver nitrate staining, but it's a starting point.

Your image from above shows very little tails. Are these control or treated cells?

Pablo, I appreciate very much your help!

I'll improve this technique, it will be much easier with this article you shared.

Best regards,

Giuseppe.

Look out for a software that lets you define the borders of the comet head and tail manually. A software like that from Andor should do this, but also other comet software: There is a Linklist at http://www.cellula.de/comet/imganl.php .

Hello Pablo, your welcome!

Hello Giuseppe: Also, your cell density is yet too high to measure, you will see that when you start measuring. Your comets will interfere with each other. Moreover, the dark spots or residues in your preparation will interfere if you measure automatically or if you try to measure the amount of DNA. Try to get a "cleaner" preparation...

Best regards, S.

Dear Sebastian,

I greatly appreciate your reply. In fact my recent slides are much more "cleaner", thanks to the help of others researchers that participate this topic. Now I can measure the comets with CASP.

Best regards,

Giuseppe.

Is OpenComet software work for comet assay image analysis?

Thanks for the suggestions... Please can you suggest me how to change settings if CSAP software is unable to detect the head of the comet correctly? Is there a macro for this?

Thanks

Best Regards

Ragavendrasamy

Ragavendrasamy, in CASP you can change settings in "options" > "adjust".

If even altering the parameters the software does not detect any comet, you image might have an issue of the direction of the comet's tail.

In the software specifications, it only detect comets that have the head in the left side and the tail formed to the right. If the image is on the opposed side, it does not detect any comet.

Other issue might be the background of your images...

Hope it helped you.

Best regards,

Giuseppe PS

How do I adjust CASP settings so that it can measure comets that are mostly tail? Right now the program thinks the middle of the tail is the head.

Thank you in advance,

A. Lin

https://www.instem.com/solutions/genetic-toxicology/comet-assay.php

Hi, Apologies for a late answer, I saw your question whilst looking for something myself. We used to use CometScore, which is free and does a good job.

http://rexhoover.com/index.php?id=cometscore

Best wishes,

Gareth Edwards.

thanks a lot everyone