Hi, every one..
I wasn't to quantify the DNA damage of the macrophage infected intracellular pathogen's DNA damage. is there any methods
Not clear question, you want to look for macrophage's DNA or pathogen's DNA?
DDR by mass spec here https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4046014/
DDR by confocal here http://www.bdbiosciences.com/documents/Bioimaging_AppNote_DNADamage.pdf
Some LC here https://www.ncbi.nlm.nih.gov/pmc/articles/PMC94673/
It depends on what are you looking for. Set that first, then it's easy to do a documentation to evaluate options
Hi, Doinita,
I want to look at Pathogen's DNA damage which is residing inside the macrophage.
If I isolate DNA from infected macrophage I will get DNA from macrophage as well as pathogen. Is there any technique by which I can only quantify the DNA damage of only pathogen but not the host cell.
So you are looking for a separation method first. First identify your sample type (blood, tissue, other biological material), then be more specific with the term "pathogen" i.e. is it a bacteria or a virus or a fungus, then look for a commercial kit which is intended for that purpose.
Once you isolate your pathogen DNA you will think of the damage to quantify
Thank you Doinita,
I want to infect the macrophages in vitro and want to quantify the DNA damage of intracellular pathogen after exposing to particular treatment.
Difficult task.
What if your treatment affects cellular DNA too, in the same way? I mean it is worth to know which kind of damage your treatment will generate, maybe affecting exactly those regions that make the difference between a pathogen DNA and it's host DNA.
If you know the expected mechanism of the treatment in damaging the DNA, am thinking now to another option. Doing a PCR with extracted total DNA with primers specific for genomic DNA, and another with primers specific for the pathogen. Before the treatment and after the treatment. Compare the two. But obviously I am missing the required elements to think more precisely, it is just a very general idea.
Good luck with it.
I agree with you.. my treatment causes nonspecific damages to both host and parasite. I am presuming that it causes more damage to pathogen's DNA. Since it is non-specific DNA damage I can't use specific primer to screen'. Therefore, isolating only parasite's DNA may be better option.
Why not to make it a comparative analysis. Keep uninfected macrophage cells from the same batch as control during drug treatment. After treatment, the difference between the DNA damage will be for the infected pathogen.
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