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We routinely use Delta treated Nunc plasticware which provides for strong adherence of Sf9 and Sf21 cells. Plasticware from other suppliers, in particular from Corning, can also be used. Insect cells growth from sparse monolayers is impeded in vent/open cap flasks and in Petri dishes that allow air exchange. In our experiments, insect cells seeded at 10% confluency or below can grow to 100% confluency in vent/close flasks, but not in vent/open flasks, Petri dishes or plates. We always use vent/close cap flasks and screw the caps tightly to seal the flask interior from the outside environment. However, performance of cells in over 50% confluent monolayers is similar in sealed and non-sealed plasticware. Therefore, semi- and sub- confluent monolayers are routinely used in Petri dishes and plates for transfections and for protein expression experiments. As a general rule, at least 40-50% confluent monolayers of Sf9 or Sf21 cells grown on TC-100 medium supplemented with heat inactivated 10% FBS should be seeded in Petri dishes, 6-well, 24-well or 96-well plates.
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