You can only run one "main" instance of Fiddler because otherwise you run into myriad problems like port contention, proxy chain confusion, settings synchronization, etc, etc.
However, there are many ways to achieve the goal of isolating traffic even within a single instance; you can use FiddlerScript to render traffic from different clients in different colors, with different icons, etc.
Alternatively, you can click File > New Viewer to create a new "Viewer" instance of Fiddler. In that new instance, in the QuickExec box below the Web Sessions list, type !listen ####
where the number is the port you want to listen on. The Viewer instance will now act as a listening proxy without changing the system's proxy settings or attempting to overwrite any of the settings of the main instance.
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