Muscle co-contraction generates joint stiffness to improve stability and accuracy during limb movement but at the expense of higher energetic cost. However, quantification of joint stiffness is difficult using either experimental or computational means. In contrast, quantification of muscle co-contraction using an EMG-based Co-Contraction Index (CCI) is easier and may offer an alternative for estimating joint stiffness. This study investigated the feasibility of using two common CCI’s to approximate lower limb joint stiffness trends during gait. Calibrated EMG-driven lower extremity musculoskeletal models constructed for two individuals post-stroke were used to generate the quantities required for CCI calculations and model-based estimation of joint stiffness. CCIs were calculated for various combinations of antagonist muscle pairs based on two common CCI formulations: Rudolph et al. (2000) (CCI1) and Falconer and Winter (1985) (CCI2). CCI1 measures antagonist muscle activation relative to not only total activation of agonist plus antagonist muscles but also agonist muscle activation, while CCI2 measures antagonist muscle activation relative to only total muscle activation. We computed the correlation between these two CCIs and model-based estimates of sagittal plane joint stiffness for the hip, knee, and ankle of both legs. Although we observed moderate to strong correlations between some CCI formulations and corresponding joint stiffness, these associations were highly dependent on the methodological choices made for CCI computation. Specifically, we found that: (1) CCI1 was generally more correlated with joint stiffness than was CCI2, (2) CCI calculation using EMG signals with calibrated electromechanical delay generally yielded the best correlations with joint stiffness, and (3) choice of antagonist muscle pairs significantly influenced CCI correlation with joint stiffness. By providing guidance on how methodological choices influence CCI correlation with joint stiffness trends, this study may facilitate a simpler alternate approach for studying joint stiffness during human movement.