Objective To develop a set of stretch strain cell loading unit with proprietary intellectual property rights so as to provide a necessary tool for the research of cellular mechanics. Methods Based on the deformation theory of round substrates, the stretch strain cell loading unit could apply the stretch strain to cultured cells in vitro by utilizing digital measuring controlling system and computer software. MTT colorimetric test was adopted to evaluate the ability of human periodontal ligament cells (HPDLCs) to attach and grow on the silastic membrane. The cultured HPDLCs were loaded with 1%, 10% and 20% stretch strain for 0.5, 1 and 24 h, respectively, by the loading unit, and an inverted phase contrast microscope was used to observe the morphology and realignment of HDPLCS. Results The stretch strain cell loading unit could apply the stretch strain of different magnitude, frequency and duration to cultured cells in vitro, with the advantage of a large range strain output, high precision, convenient operation and clear visual monitoring. When cells were seeded for 1, 2, 4, 7, 8 d, respectively, there was no statistical difference between the MTT absorbance value of silastic membrane group and that of control group (P>0.05), showing the good ability of cultured cells to attach and grow on the silastic membrane. After HPDLCs were loaded with 10% and 20% strain for 24 h, the morphology and realignment of HPDLCs changed, and cells became spindle shaped, being parallel to each other with long axis perpendicular to the stretch vector. Conclusions The stretch strain cell loading unit is capable of loading the stretch strain to cultured cells effectively and provides a necessary method for cytomechanical research.