Ha! Yes, ever since I was a kid.
On my first day teaching an Algebra class, my students made a sign for the door that said "Algebra I: MR teacher". Once when I was buying bras, I went to the fitting room clerk to get a room unlocked and she walked straight to the door marked "Gentlemen" and opened it up for me. I studied abroad in Japan for a semester and frequently had groups of three or so high-school girls whispering about me on the train and sneaking glances my way. Or else straight up talking about me, thinking I couldn't understand Japanese. (Enlightening them was often quite entertaining.) One Japanese guy asked me whether I was a boy or a girl before he started hitting on me. My family went to the dentist here in the States and the nurse asked my teenage sister if I was her boyfriend. (When she finished with the tooth scraping, my sister pointed out her mistake. We laughed about it in the car afterwards.)
I'm very active, and I like my hair short and my jeans not too fitted. Usually, I laugh when I get mistaken, and (with the exception of a Russian lady and an old man in a bookstore, and maybe the Japanese guy) everyone who talks to me for more than two seconds knows I'm a girl.