Hello,
I am trying to measure the cytokine storm markers following SARS-Cov-2 viral infection of lung cells and macrophages under different conditions. However, finding a safe way to deactivate the virus while preserving the markers is not straightforward to find in literature, any suggestions?
I would think you need to define which stage of infection you want to assess the cytokine profile. RNAi might work against one or more key packaging genes, or ribosome inhibitory proteins/compound and proceed with extraction of cytokines/total proteins. But the former can be designed at any stage of your preference, while the latter would halt all protein synthesis including cytokines. A double chambered culture vessel might help to try both approaches to see multistep cytokine events. Hope this helps. Alternatively you can transfect an attenuated virus but the immune response might be different than actual, or you can add one of the reported virus killing compounds to the medium such as GC376.
I am testing for drugs that attenuate the cytokine storm, so I really need to have a full blown infection, I will be using a co-culture method using hydrogel to separate the two cell cultures, but I'm stuck at finding the right method to deactivate the virus before proceeding with the detection. One way to avoid this I believe is to use Elisa kits, if I can find a suitable one, that allows me to detect certain markers at the cell culture lab, but I'm not sure that would be available I have to check.
You mean to detect cytokines by ELISA? Because ELISA cannot deactivate the virus! You may add antibodies to medium to illicit an immune response in your cells. It depends really if you want to see the full infective cycle of the virus up to release of new viruses.RNAi should help you control the infective cycle stages in the lung cells.
yes of course, the Elisa is for cytokine detection, not the virus. I have to admit I didn't consider the infective cycle, I was planning towards 1h incubation time of lung cells with the virus then another one with the macrophages.
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