I am using rats ovarian tissue in processing Comet assay and I follow the COMPAC-50 high throughput comet electrophoresis system protocol by Cleaver Scientific Ltd.DNA dye that used was Vista green. I have problem in getting very clear image when I am using 100X power.
Dear Ali Saeed Al-Chalabi,
How are the images of lower magnification (lesser than 100x) ?. Are you able to visualize cells/comets clearly in lower magnification ? . Might be there is less cell density in the slides.
it looks like there isnt any DNA damage , please sheer the DNA with sonicator and optimize the protocol. you can take sybr green I dye as well. then take that much amount of cells which will generate the same DNA. BY the way are you performing electrophoresis or not? if not please do.
Dear Samrat Das
thanks for your feedback, actually i used 40X power
It seems like the concentration of your drug is too high. Please try lowering it or check it first with cell viability assays like MTT, TTC or MTS. The small DNA fragments sometimes tend to diffuse in the agarose gel making it harder to analyse.
DEar valued Saadia Ijaz
Thanks for your feedback, actually I am not using drug, I am trying to see the genotoxic effect of EMF exposure on ovarian tissue.
OK! Then try lowering the duration of exposure! What is the exposure time?? Firstly try to adjust your exposure time and frequency as well. Your last image is showing obvious results of over exposure with diffuse fluorescence and there is hardly any DNA in the nuclei. Thats what I understood from your images. I hope I helped somehow!
I am testing the effect of 1800 MHz radiofrequency GSM-like ( mobile phone frequency) after daily 1 & 2 hours exposure mimic to normal phone use in human. so I understood from your comment that the EMF exposure has genotoxic effect via DNA damage showed by Comet assay, right?
Dear Harald Kunel
Thank you very much for your valued feedback.
Regards Ali
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