i used 10 microliter ethidium bromide for each loaded samples(two) on slide(i.e 20 microliter for each slide) but under the fleurosent microscope, can hardly see the halo DNA or comet.
One of the captured photos is attached.
Please see.
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Hi.
It depends on the concentration of your EtBr Stock solution. If it wasn't high enough, then it might not be visible.
Conversely, if it was too concentrated, the cells might be overstained..
However, from the picture you sent, it seems that the concentration wasn't too strong. 10ul is okay per window. Try a slightly higher conc.
Also, confirm that the gels are actually on the slide because it might be that there is no gel there.
In addition, it could also be that there are no cells on the gel. You should try and confirm all of these.
All the best.
hi temitope.
steps my work in the process of cell suspension:
1- after the exit microtube(from freezer -80C) containing liver tissues and PBS buffer, Samples are placed on ice until the ice melted
2- then samples transfer into fresh microtube and Added 200 microliter PBS and We started to cut and homogenized tissues by scissors.
3- after the homogenization, samples are vortexed once and put on ice(Due to the large pieces are deposited).
4- After a few minutes, supernatant is removed with microsampler and transfered to new microtube
5- centrifuging with 1170 rpm,7 minute at 4 C
6- removing supernatant and added 100 microliters PBS and vortexed once
7- again removed the supernatant and added 10 microliter PBS and 100 microliter low melting point agarose
8- finally, after the few times Pipetting, solution loaded on slides and covered by coverslips
Do you confirm the above steps?
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