Hi all,
I have been doing plaque assays of SARS-CoV-2 using Vero E6 cells modified to express ACE2 and TMPRSS. Cells are plated at 10^5 in one mL 24 hours before in 12 well plates. Upon staining, I notice clear circles wherein no stain uptake occurs (presumably due to cells dying there). I suspect that it is due to the cells drying out as I have been removing and adding media to 3-4 plates at a time, and I am working under a strong hood in a Cat 3. I could be mistaken by what I believe to be the cause of the problem, has anyone experienced this issue and found the resolved it? If so, what did you find helps?
Many thanks in advance
What you proposed as the reason is correct. When removing media from plates, the residual media surface tension generates a thin zone in the well centre that will cause cell damage quite quickly.
The cause of the plaque is the CPE which mean pathological changes in the cell(necrosis)which can be picked in single cell(pathological) for purification or titration.
I have being do plaque assay using fibrblast and circle of dead cell visible and can be picked or counted.
I just write my simple partciptate about plaque .thanks for whom write and comment about SARS_CoV_2.
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