I have a moderate sized eBay store, which I operate as a business, so don't always reply to messages as they're received, particularly on weekends. 

Namely because I don't want to set precendents that are not sustainable, it might sound weird, but I often am up late at night, and often it's because I'm working, but I don't want to create expectations that I can answer messages 24 hours a day, 7 days a week (not least of all because sometimes, I need a little time to stew over the reply ). 9pm is the general cut-off point for me to answer queries on weekdays, unless there's special circumstances (I have replied later than that but prefer not to unless for special customers, regulars and the like - I don't really feel that people expect instant replies, but more just taking the chance to send the message while they have it). 

On weekdays, I'll answer simple queries as they come in, and more involved queries (especially if they involve custom orders) at specific times, mostly when I have more time to respond again quickly to further enquiries and/or create a special listing, so usually after I get home from posting. Some days, if I answered all queries as they came in, nothing would actually get posted, so for me it's about effective time-management. 

On weekends, I tend to answer any messages once per day in the late afternoon, unless one is an ongoing conversation or it's genuinely urgent. It's impossible for me to have the weekend off, unfortunately, and I tend to work all day Sundays (otherwise Mondays would be a bit of a struggle), but I like to have as much of the Saturday as I can to not have to do anything selling-related.