Title says it all. When I say it, I often pause after now. Not sure if it's correct to have the comma there or not.
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If you pause after the Now, when saying Now, on with the show!, then putting in a comma indicates that pause.
It isn't right or wrong to include or omit the comma. But (IMHO), most people would probably pause after the Now (as you do), do it's probably better to include the comma.
A per def. 2 of Oxford the now you are using is to get attention to the rest of the sentence, in a conversational manner. Hence, both the examples in my link have a comma after the now.