Day 1.
Treat cells with 1X EdU from 1000x stock. . Add 0.2 ul/2 ml complete media.
Day 2.
Wash cells with 1x PBS
Fixing: add 500 ul of 4% PFA. Incubate at RT for 15 min.
Wash 1x PBS X2
Add 0.3% Triton X-100, incubate at RT FOR 10 min.
Wash 1x PBS X2
Prepare following Rn.
1x click IT buffer: 1.7 ml
CuSO4 (Component D): 80 ul
Alexa Fluor (Component B) 5ul
1x ClickIT buffer additive 200ul
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Final vol 2 ml
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Add 250 ul/well incubate at RT 30 min.
Wash cells with 100 ul of rn. rinse buffer
Add DAPI
Mount