All the time. Most people I chat with online assume I'm a gay man (I mention my husband a lot). When they learn I'm actually a female named Jay, the little light bulb goes off-- "oooooh!"
It's amusing and I don't get offended, but I also realize I am not making it easy for them-- don't exactly broadcast my sex-- my avatar on the forums is a guy and his dog. My twitter? A cat. Honestly, I would totally be fine being a married gay guy-- we'd be such a cute birding, geeking, pet parent couple.
I remember your Toad avatar as well, but I assumed you were a girl because of your protag being a girl! I don't know why I made that connection, because I read comics created by females with male protags (vice versa) and don't assume that. Maybe I came across their name in their comic credits? Interesting fun topic...