Hello everyone,

I am interested in immunostaining cell-laden hydrogel scaffolds for secreted proteins, such as VEGF. I can do both whole scaffold immunostaining and also cryosection. I typically wash my samples before a fixation step. For VEGF, I believe that it is secreted and stays soluble (Does any portion of it become immobilized in the ECM?) Is it important to wash samples after culture has ended before fixation? Should I minimize the number of washes to keep the VEGF in the scaffold?