The initiation and progression of knee and hip arthritis have been related to limb loading during ambulation. Although altered gait mechanics with unilateral lower limb loss often result in larger and more prolonged forces through the intact limb, how these forces differ with traumatic limb loss and duration of ambulation have not been well described. The purpose of this study was to determine whether biomechanical variables of joint and limb loading (external adduction moments, vertical ground reaction force loading rates, and impulses) are larger in the intact limb of servicemembers with versus without unilateral lower limb loss and whether intact limb loading differs between shorter (≤ 6 months) versus longer (≥ 2 years) durations of ambulation with a prosthesis. A retrospective review was conducted of all clinical and research gait evaluations performed in the biomechanics laboratory at Walter Reed Army Medical Center and Walter Reed National Military Medical Center between January 2008 and December 2012. Biomechanical data meeting all inclusion and exclusion criteria were obtained for 32 individuals with unilateral transtibial limb loss, 49 with unilateral transfemoral limb loss, and 28 without limb loss. Individuals with unilateral lower limb loss were separated by their experience ambulating with a prosthesis at the time of the gait collection, ≤ 6 months or ≥ 2 years, to determine the effect of duration of ambulation with a prosthesis. Intact limb mean and peak vertical ground reaction force loading rates (median [range; 95% confidence interval]) were larger for transtibial subjects with ≤ 6 months of experience ambulating with a prosthesis versus control subjects (mean: 12.13 body weight [BW]/s [4.45-16.79; 10.18-12.81] versus 9.03 BW/s [4.64-14.47; 8.26-9.74]; effect size [ES] = 0.40; p = 0.003; and peak: 17.23 BW/s [6.58-25.25; 15.46-19.01] versus 13.60 BW/s [9.82-19.51; 12.98-15.05]; ES = 0.43; p = 0.001), respectively. Intact limb mean and peak vertical ground reaction force loading rates were also larger in subjects with transfemoral limb loss with ≤ 6 months and ≥ 2 years of experience ambulating with a prosthesis versus control subjects (mean: 12.67 BW/s [5.88-18.15; 11.06-14.47] and 12.59 BW/s [8.08-17.39; 11.83-13.68] versus 9.03 BW/s [4.64-14.47; 8.26-9.74]; ES ≥ 0.53; p < 0.001; peak: 19.82 BW/s [11.93-29.43; 18.35-23.05] and 21.33 BW/s [16.68-36.69; 20.66-24.26] versus 13.60 BW/s [9.82-19.51; 12.98-15.05]; ES ≥ 0.68; p < 0.001, respectively). Similarly, intact limb vertical ground reaction force impulses (0.63 BW·s [0.53-0.81; 0.67-0.69] and 0.62 BW·s [0.55-0.74; 0.60-0.63] versus 0.57 BW·s [0.50-0.66; 0.55-0.58]; ES ≥ 0.53, p < 0.001) were also larger among both groups of transfemoral subjects versus control subjects, respectively. Limb loading variables were not statistically different between times ambulating with a prosthesis within groups with transtibial or transfemoral limb loss. Larger intact limb loading in individuals with traumatic transtibial loss were only noted early in the rehabilitation process, but these variables were present early and late in the rehabilitation process for those with transfemoral limb loss. Such evidence suggests an increased risk for early onset and progression of arthritis in the intact limb, especially in those with transfemoral limb loss. Interventions should focus on correcting modifiable gait mechanics associated with arthritis, particularly among individuals with transfemoral limb loss, to potentially mitigate the development and progression in this population.