The invention belongs to the technical field of robots, and particularly relates to a pre-tightening force adjusting structure, a chassis suspension assembly and a robot, wherein the pre-tightening force adjusting structure comprises a connecting rod, an elastic element, a guiding column and a fixing element, wherein an installing hole is formed in the connecting rod; the elastic element is installed in the installing hole; the first end of the guiding column penetrates through the installing hole and the elastic element extends to the outer of an orifice of the installing hole; and the fixingelement is arranged at the first end of the guiding column. When the second end of the guiding column is fixed on a structural element, the relative position between the fixing element and the guiding column is adjusted to adjust the scalability of the elastic element. According to the pre-tightening force adjusting structure, the structure is simple, the scalability of the elastic element is adjustable, and different pre-tightening forces can be applied to the elastic element according to different application scenarios.