Hello all,
I've recently been exploring the use of ThT as a membrane potential dye, I was hoping to ideally fix cells after I have treated the cells with ThT, however I have not found any protocols stating this is possible.
Has anyone done something similar to this?
Regards,
Update: Due to the lack of response I decided to experiment.
Using a density of 5x10^4 HeLa cells in a 12 well plate, cells were incubated at 37C in 5%CO2 for 24 hours to allow for attachment and partial proliferation.
Media was removed, cells washed twice with PBS and various concentrations of ThT (10-1mM) in HBSS was added and incubated at room temperature for 30 minutes. Cells were then examined in 3 stages, directly after incubation without removing media, post washing and after fixing with 4% formaldehyde.
Cells observed in media were found to comfortably fluoresce down to 1mM at which point signal started to decrease, similar results were found in post washing samples.
Cells that had been washed and fixed demonstrated significant loss of signal with only samples treated with a 10mM concentration providing adequate fluorescence.
I hope this helps anyone who comes across this question, regards.
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