I am trying to measure secreted cytokines TNF-alpha and IL-1beta from RAW264.7 cell lines after LPS stimutaion. I am unsure whether the cells should be serum starved for performing ELISA from the cell culture media. Please provide your comments based on your experience.
Serum starving of cells, depending upon the length of starvation, is likely to reduce protein synthesis. Therefore, upon stimulation with LPS, one is likely to see minimal induction of cytokines. To perform ELISA for cytokines, one does not need starvation as long as a control is used.
Serum starving of cells, depending upon the length of starvation, is likely to reduce protein synthesis. Therefore, upon stimulation with LPS, one is likely to see minimal induction of cytokines. To perform ELISA for cytokines, one does not need starvation as long as a control is used.
Thank you, Francis Corazza!
I am not sure what is IPS binding protein in the serum is. Could you please explain the details of it. Please elaborate what its mechanism is and why it is necessary.
Thank you.
Thank you Sir, Divaker Choubey. It is helpful.
Could you also help me determine the cell density which is required to obtain measurable levels of the cytokines by the ELISA kit? I was planning to use 20,000cells /cm2 in a 6well plate and add 2ml of media. Is this cell density and the media volume fine to mesaure the cytokines by ELISA? I am concered if the cytokines will be diluted a lot due to low cell number and 2ml of media volume.
Kindly share your experiences regarding it
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