The present invention is to provide a dental floss holder containing a body that includes a fixing portion, which is provided with at least a recess disposed inward so as to constitute a neck for holding the dental floss and is provided with at least a slot for receiving the dental floss in position. Thus a user can operate the dental floss fixed on the body in cooperation with the other end of the dental floss held by fingers (or fixed on another dental floss holder) to easily clean the slits between teeth deep inside the oral cavity.