I would like to perform an ELISA from the cell culture supernatant of immortalized human hepatocytes (IHH) cells.
When I attempted the ELISA for a first go, the apoB concentration was on the lower end of the standard curve (~14 ng/mL). Would you have any recommendations on how I should proceed to concentrate my cell culture supernatants for ELISA?
Thank you for your help.
Hey Alessandro,
we had the same problem with our mouse embryonic fibroblasts. The easiest solution you could try is to seed as much cells as the well can bear and keep your medium volume (so your supernatant) as low as possible (of course the cells should be happy and not dry or hunger out).
The other possibility is (since I do not know your stimulus, which should induce cytokine production and secretion in your cells) that your stimulus is either not inducing cytokine production or the concentration is not right.
Do you have any further information on that?
You can also combine these two possibilities (stimulating many cells in a small volume with a lots of stimulus).
All the best,
Marc
You can try with PEG
Article Protein precipitation with polyethylene glycol
You could simply load your supernatant onto a Millipore spin concentrator depending of the size of your protein you choose the appropriate cut-off
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