"Halo" or edge-ring artifact around dishes (Colony Formation Assay)

Question

Accepted Answer

A ring of stain appears around the dish edge; cells aggregate at the periphery due to evaporation / meniscus effects. Common causes: 1) Fixation was inadequate 2) Wash water was poured in too hard and washed cells to the edge 3) Medium evaporation produced an edge effect during long incubation Solutions: 1) Fix adequately (15 – 20 min in 4 % paraformaldehyde) 2) Wash gently — submerge or pipette slowly along the wall 3) Cover and seal dishes during the long incubation; maintain incubator humidity > 90 %