Objective To describe the effects of axial loading, small-scale parameter, bioliquid density in a microtubule (MT) and constrained stiffness of surrounding biomedium on the coupling vibration frequency in the bioliquid-filled MT, and to provide references for ultrasonic inspection on nano-MT under axial loading and the clinical application of biological medicine. Methods The non-local elastic theory was utilized to describe the nano-scale characteristics of the MT, and the analytic solutions to the coupling vibration frequency of the bioliquid-filled MT under axial loading were given. Results The axial loading exerted on the bioliquid-filled MT made the couple vibration frequency drop rapidly, and as the small-scale effect increased, the couple vibration frequency of the bioliquid-filled MT was gradually decreased. The effect of axial loading on the couple vibration frequency of the bioliquid-filled MT was larger than that of the small-scale parameter. Conclusions When the density of bioliquid in MT increases, the first order frequency of the bioliquid-filled MT embedded in biomedium is decreased; when the initial axial loading exerted on the bioliquid-filled MT increases, the effect of bioliquid density in MT on the first order frequency of bioliquid-filled MT is reduced gradually.