i am trying to grow TTV on 293T, Jurkat, and A549 cell lines for a prolonged period. As the replication mechanism of this virus is not fully understood, I want to check what receptors on these cell lines are facilitating the attachment of the virus.
It would be best to pull down the viral capsid using immunoprecipitation in non-reducing conditions after infecting your target cells. After confirming the successful pulldown by detecting loads of capsid protein in the pulldown, you can then perform a mass spectrometry analysis of everything that is contained in the pulldown. Then, you analyze the detected proteins in the pulldown to choose the candidates for the knockout. If you successfully knock out the binding receptor in your cells the viral infectivity will be dramatically reduced.
Thank you so much for your answer. Can you please provide me with the protocol?
First, you need an antibody that binds well to the capsid of your virus and confirms that it works by western blotting. The next step is to follow one immunoprecipitation protocol available online. They are usually tailored to a specific kit or set of reagents. For example, you can use Pierce Co-IP kit. https://www.thermofisher.com/order/catalog/product/26149
After successfully confirming the pulldown of your capsids, you can communicate with your institution's Mass Spec core or contact any service for a fee mass spec centers available in your location.
thank you so much for your suggestion. i discussed this with my supervisor. do i need anti-TTV antibody that can bind to capsid of TTV ?
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