I am currently working with Sf9 cells and have been testing various growth conditions. Specifically, I’ve tried using Insectagro SF9 medium supplemented with different concentrations of FBS (3%, 5%, and 10%) along with 1% Penicillin/Streptomycin, as well as serum-free conditions. I've been culturing the cells at 27.5°C in an incubator.

Initially, the cells appear healthy and grow well through the first passage, as seen in the image labeled "Picture 1." However, after about a week, they start to undergo unusual morphological changes, resembling crystalline and tadpole-like shapes (see attached pictures for reference). I’m wondering if anyone has encountered similar issues or has suggestions on how to maintain stable cell morphology over longer passages. Any insights into potential causes or adjustments to the media or environmental conditions would be greatly appreciated!

Regards,

Muhammad