Please can someone indicate the typical dilution factor used for cell culture supernatants in the detection of cytokines. The cells used are mouse and human macrophages (stimulated and unstimulated)
For LPS activated RAW264.7 macrophages we dilute supernatants 1/40 for IL-6 ELISA. We assay NO on the neat supernatant.
For human whole blood cell activation with LPS we dilute supernatants 1/40 for IL-6 ELISA.
For PHA and ConA activated whole blood cultures we analyze the neat supernatants for IL-10 and IFNg.
Let me know if you require more info.
For LPS activated RAW264.7 macrophages we dilute supernatants 1/40 for IL-6 ELISA. We assay NO on the neat supernatant.
For human whole blood cell activation with LPS we dilute supernatants 1/40 for IL-6 ELISA.
For PHA and ConA activated whole blood cultures we analyze the neat supernatants for IL-10 and IFNg.
Let me know if you require more info.
Your response has helped me so much you have no idea! I have been looking all day at the literature to get answers and no one indicates their dilution factors!!! What about TNF and IL-12 and 1? And Do you know anything about dilution factors for RANTES and MCP-1? Thank you a million times!!
Hi Gina, I only do the above cytokines. Check with Prof Pat Bouic. The people in the Microbiology/Immunology Department on 10th floor in Tygerberg Hospital will have his contact details. Alternatively try Dr Tandeka Magcwebaba in the Biochemistry Department on Stellenbosch main campus. Prof Carine Smith in the Physiology Department on Stellenbosch main campus may be another source close to you. All of the mentioned do cytokine ELISAs. I normally start with a dilution curve of my supernatants. You can message me if you need further contact details.
Thank you very very much!! I will contact them ASAP. You have been a great help!
Hi Gina, there is a link on the attached document that will lead you to a website where they give the dilution factor for each cytokine. Hope it helps
Hi Gina, we use the culture supernatant as it is (nit) without any dilutions and it gives us proper results.
@Vishal, I dont think this will work with my samples as I have stimulated the cells with M.tb. Thus I expect the synthesis of cytokines to be increased substantially. qPCR indicates this.
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