we are having a common problem in the lab with recombinant secreted protein expressed in insect cells (both Sf9 and Hi5).
1) protein does not bind when Ni-NTA or Talon is directly added to the clarified supernatant
2) 4L conditioned media preciptation with AMSO4 results in fluffy precipitates.
3) apparently, there is detergent in the media which is added to prevent shear stress to the cells during culturing, which affects binding to NiNTA. pHing the media to 8 crashes the detergent out - but the further clarified supernatant also shows poor to no binding.
What is the best way to purify secreted protein in media in insect cells.
Suggestions would be greatly appreciate!