Hi all,
So I want to see if my protein is secreted into the media of my cultured cells and wondering if the correct protocol would be to just collect the media and adding sample buffer to it and just running it on the gel?
Please if you have tried it before or have a better protocol please share it with us?
Many thanks!
Yes, See my papers
Figure 4D of
Article Endocytosis and serpentine filopodia drive blebbishield-medi...
and
Figure 8c of
Article Blebbishields, the Emergency Program for Cancer Stem Cells: ...
See methods: Analysis of VEGF secretion in second paper.
Good luck..!!
Jinesh.
Yes this is one classical approach to determine if your protein of interest is expressed and secreted properly and compare this to the purified protein. If your protein of interest is not secreted at high yield it is sometime necessary to concentrate your conditioned culture media. It is also interesting to have as control your cell pellet (well washed) to see if your protein is expressed.
You have to concentrate your serum sample using commercially available protein concentrater , as the increased dilution may result in false negative or too dim results.
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