Hello I work in sw620, sw480 and HT 29 cell lines and I did a migration assay for 29 H using Dapi . the problem is that i didn't get enough fluoresence

I put a 200000 cell in 200µl free medium within insert (8µm pores) and I put 800 µl of medium containg 10% SVF.

this is my protocol :

fixing the cells with methanol for 20 min (200µl in inserts) and (800µl in wells)

washing cells with water

adding triton X with 1% concenrtration (200µl in inserts) and (800µl in wells)

washing cells with PBS

Removing cells in the upper side of the membrane inserts

cutting the membrane

adding a drop of dapi in the blade

this the file for watching