Hello. I plan to do a simple immunofluorescence procedure with HT-22 cells. I plan to transfect the cells and fix them. I do not believe I will have time to come in on the weekend. Is it possible to store my cells after fixation? If so, should I dehydrate them using ethanol? Are there any other recommendations? After fixing with PFA the next step is to wash and incubate with Triton 100x. In addition, will this disrupt my signal or my cells?? Thank you
Hi,
You can fix your cells with PFA (room temperature) or ice-cold methanol (4 C) for 15 minutes, then rinse with PBS 2-3 times. Cells can be left in PBS in the fridge and this won't affect fluorescence. When you are ready you can resume the experiment starting with the permeabilization/blocking step (with triton and BSA).
Worked all the time for me
Hope it helps
I did this protocol many times. After the fixation and the washes, I usually store cells in PBS1X at +4°C for a maximum of 1 week.
You can use the triton in the permeabilization step and/or in the blocking step but it depends on the antibody you will use.
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