not a simple yes or no question
normally no. all the 2 and 3ds systems recognize cards formatted with FAT32. Because this card is bigger than 32 gigs, most formatting programs will only let you format it with EXFAT which 2 and 3ds systems won't recognize.
This card will work on a 2 or 3ds. I have one of these in my new 3ds. Unfortunately you either have to purchase or hunt down a formatting program that will allow you to format the card in FAT32.