I've been searching for any studies relating to the study of cell culture supernatant and the potential biomarkers that may present within unprocessed media. I keep running into paywalls, dead ends or unrelated topics. Can anyone point me to any articles or reviews remotely related to this area?
Thanks for the response Cristian; I have yet to chose a specific cell line, given I plan to assess toxicological factors of xenobiotics, I will most likely be using a hepatocyte line. I'm currently just trying to establish some background reading in supernatant analysis to decide which media/ cell line would be easiest to monitor in real-time, hence biomarker monitoring.
Screen the culture supernatants for lactate dehydrogenase (LDH). Can be done with an enzymatic assay kit (several available commercially and relatively cheap). This will tell you if the sample is toxic. Upon exposure to toxins the cell membrane leaches cytoplasmic proteins such as LDH. You can also analyse heat shock protein in the medium. Will give you an indication of stress. Also check for nitric oxide (NO). Will indicate inflammation. For NO assays make sure you use DMEM and not RPMI as RPMI give high backgrounds.
Forgot to expand on heat shock protein: ELISA kits are available for these eg. HSP70.
For NO: Griess assay works very well and is cheap.
For the Greiss assay, remember the half life of nitrite in blood is very short (a matter of seconds - converts to nitrate), therefore you have to use nitrate reductase to convert nitrate to nitrite in order for the Griess reaction to work. I presume the same effect will occur in various in vitro systems, to a varying degree. So a general kit from Promega will be good, you will need the Nitrate/Nitrite Colorimetric Assay Kit from Cayman Chemicals. Here is article on the Determination of Nitric Oxide via Measurement of
Nitrite and Nitrate in Culture Media (http://lifescience.roche.com/wcsstore/RASCatalogAssetStore/Articles/BIOCHEMICA_2_95_p22.pdf). Also need to remember to avoid media with phenol red and hugh nitrate e.g. RPMI 1640.
Have you thought of electrochemistry? Have a look at:
http://eprints.uwe.ac.uk/17914/
If so, I have some other papers you may be interested by.
Kevin
Hey Kevin, thanks for the kind offer. Conincidentally, my PhD is based around the development of electrochemical sensors and their use in cell media, so your work, Dr Hart's and Dr Pemberton's have been a great influence. This question in particular was proposed when my concept was at quite a juvenile stage but fortunately it's developed into looking at ROS and associated reactive species. Thanks once again.
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