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Dear Colleagues,
I have some problems with mounting insect cells (Sf9). I grow my cells on coverslips (13 mm round) and infect them with baculoviruses. After 48 h the cells are fixed in 4% paraformaldehyde. At this stage the cells looks fine, but when I mount the cover slips the cells shrink immediately and become sharp-edged instead of nicely round cells. The cells are unusable for (fluorescence) microscopy. I have used different mounting solutions (VECTASHIELD HardSet Mounting Medium, VECTASHIELD Mounting Medium, Dako Fluorescence Mounting Medium, PBS:glycerol (1:9)) but they all give the same problem. I have tried to coat the coverslips first with poly-L-lysine or poly-L-ornithine, but this had no effect. When I mount them in some PBS they look fine, but this is not very workable. When I use Hela or Soas-2 cells they look fine, so it is something specific for Sf9 cells. Is there someone who has experience with this type of cells? I am trying to detect GFP, TagRFP and Hoechst.
With kind regards,
Marcel Westenberg
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