This study aimed to prepare an anti-Vascular cell adhesion protein 1 (VCAM-1) nanoscale ultrasound microbubble contrast agent using the hyperbranched self-assembly method for the molecular imaging diagnosis of atherosclerotic vulnerable plaques in rabbits. Twenty-five rabbits with carotid atherosclerosis were randomly divided into 5 groups, and the ear vein was injected with agents as follows: Groups A and B: nanoscale ultrasound microbubble contrast agent with and without anti-VCAM-1 agent; Groups C and D: SonoVue ultrasonic microbubble contrast agent, with and without anti-VCAM-1 agent; Control group: saline. The molecular imaging diagnosis of the atherosclerotic plaque, involved the examination of its vulnerability in the rabbit carotid artery was performed using the contrast ultrasound mode. The arrival and peaking time of the anti-VCAM-1 nanoscale ultrasound microbubble contrast agent (Group A) for plaque occurred earlier than those of the other groups (p 0.05), and with it, the plaque showed the strongest enhancement (p 0.05), followed by the SonoVue ultrasound microbubble contrast agent with anti-VCAM-1 group (Group C) and the self-made nanoscale ultrasound microbubble contrast agent group (Group B). No development was observed in the plaques of the SonoVue ultrasound microbubble contrast agent group and the control group. The anti-VCAM-1 nanoscale ultrasonic microbubble contrast agent, prepared using the self-assembly method, can facilitate the development effect of the carotid atherosclerotic vulnerable plaque, providing a basis for the molecular imaging diagnosis of carotid atherosclerotic vulnerable plaques.