Blood Flow Restriction (BFR) training has gained growing attention in musculoskeletal rehabilitation. Effects of resistance training with and without BFR on lower-limb muscle activation were analyzed in a crossover study including 8 subjects (27.5 ± 4.8 years) engaged in regular CrossFit^® training. Two 48 h-separated sessions (high bar back squat), with BFR (80% of individualized arterial occlusion pressure) and without BFR, were performed in a randomized order. Muscle activation (vastus lateralis, vastus medialis (VM), gluteus maximus (GM) and biceps femoris) was electromyographically recorded. The average electromyographic activity of the first four repetitions of the first set (T1), and the last four of the final set (T2) were considered, and intra- and inter-group comparisons were calculated. Pain (VAS) and fatigue (Borg scale) were recorded. Significant differences between BFR and non-BFR conditions were observed only for T1 peak BF activation values. Within the BFR experiments, no significant differences in EMG activation were detected between T1 and T2. In contrast, within the non-BFR experiments, T1 values were statistically significant higher (vs. T2) for mean VL, mean VM, peak VM, mean GM and peak GM. Median pain perception was 3.5 (0.75–6.75) and 4.0 (1.25–6.0), and fatigue perception was 8.0 (7.0–8.0) and 7.0 (7.0–8.0), respectively, with and without BFR. In this exploratory study, BFR maintained EMG activation (VM and GM) and suggests a strategy to minimize mechanical load while preserving muscle activation.