Quadriceps muscle weakness is a commonly reported issue post anterior cruciate ligament reconstruction (ACLR), with minimal information related to skeletal muscle morphology following surgery. The purpose is to examine the morphological and functional differences in the vastus lateralis muscle from patient's ACLR and contralateral leg. Three physically active ACLR participants were recruited and secured to a dynamometer to perform maximal voluntary isometric knee extension contractions (MVIC) of the ACLR and contralateral limb. Muscle biopsies of the ACLR and contralateral vastus lateralis were performed, then sectioned, and stained for myosin isoforms to determine fiber type. Confocal images were acquired, and ImageJ software was used to determine the fiber type and cross-sectional area (CSA). There was a significant reduction in CSA of the type IIa and type IIx muscle fiber cells between healthy (IIa: 7,718 ± 1,295 µm2; IIx; 5,800 ± 601 µm2) and ACLR legs (IIa: 4,139 ± 709 µm2; IIx: 3,708 ± 618 µm2) (p < 0.05), while there was no significant difference in knee extension MVIC torque between legs (healthy limb: 2.42 ± 0.52 Nm/kg; ACLR limb: 2.05 ± 0.24 Nm/kg, p = 0.11). The reduction in the cross-sectional area of the ACLR type II fibers could impair function and increase secondary injury risk.