This study details the concept, design, and mechanical testing results of a novel dual-stiffness ankle-foot orthosis (DS-AFO). The DS-AFO utilizes two separate stiffness elements (rear struts) to yield an AFO with low stiffness properties about the ankle in the sagittal plane at small dorsiflexion angles, and higher stiffness at larger dorsiflexion angles. The motivation behind this DS-AFO follows from the existence of similar moment-angle (stiffness) properties of the healthy human ankle during walking, referred to as dual-stiffness natural ankle quasi-stiffness (DS-NAS). Crucial to the design of the DS-AFO is the ability to adjust both the stiffness and the dorsiflexion angle at which the net stiffness increases, referred to as the activation angle. Three different DS-AFO stiffness configurations were tested, each with three different activation angles, along with a standard single strut/stiffness AFO configuration. The DS-AFO was able to achieve distinct regions of low and high stiffness at every configuration. Additionally, altering the activation angle by ±1 deg generally did not result in different stiffness properties. This work is a step forward in AFOs with complex stiffness properties that can better approximate the mechanics of a healthy human ankle.