Botulinum toxin type A (BTX-A) injections in masseter muscle can alleviate muscle tightness and aching pain caused by idiopathic masticatory myalgia, a subform of the myofascial pain syndrome. Yet the injection procedure (number, amount) is currently empirical. In this ex vivo study, we determined the feasibility of using contrast-free ultrasound imaging to visualize the short-term injectate propagation. Ultrasound annotations of BTX-A injectate spread in N = 12 porcine masseter muscles were compared with the histopathology of the excised masseter. BTX-A presence was automatically detected in the ultrasound cine by: compensating tissue motion and deformation during injection with a novel spatiotemporal filtering (SF) algorithm, and by imaging tissue swelling strains with strain elastography (SE). BTX-A injectate introduced 6.5% (standard deviation = 5.0%) echogenicity contrast and 13.9% (standard deviation = 3.7%) tissue swelling strain. Muscle fasciae were a border for BTX-A distribution. The SF algorithm achieved significantly higher noise rejection (contrast-to-noise ratio = 4.63) than SE (2.56, p = 0.01), and state-of-the-art 2-D digital image correlation (1.81, p < 0.001) and direct image subtraction (1.29, p < 0.001) methods. Histopathology agreed well with ultrasound (Dice coefficient = 0.48), with deviations mainly explained by the three-dimensional inhomogeneous distribution of BTX-A. Preliminary in vivo patient results indicated that SF and SE discard artifactual BTX-A detection outside the injection region. The proposed methods contribute to objectivize ultrasound-guided injections, with additional applications, for instance, to monitor injectate spread of local anesthetics.