Cell-substrate interaction is implicated in many physiological processes. Dynamical monitoring of cellular tractions on substrate is critical in investigating a variety of cell functions such as contraction, migration, and invasion. On account of the inherent ill-posed property as an inverse problem, cellular traction recovery is essentially sensitive to substrate displacement noise and thus likely produces unstable results. Therefore, some additional constraints must be applied to obtain a reliable traction estimate. By integrating the classical Boussinesq solution over a small rectangular area element, we obtain a new analytical solution to express the relation between tangential tractions and induced substrate displacements, and then form an alternative discrete Green’s function matrix to set up a new framework of cellular force reconstruction. Deformation images of flexible substrate actuated by a single cardiac myocyte are processed by digital image correlation technique and the displacement data are sampled with a regular mesh to obtain cellular tractions by the proposed solution. Numerical simulations indicate that the 2-norm condition number of the improved coefficient matrix typically does not exceed the order of 100 for actual computation of traction recovery, and that the traction reconstruction is less sensitive to the shift or subdivision of the data sampling grid. The noise amplification arising from ill-posed inverse problem can be restrained and the stability of inverse solution is improved so that regularization operations become less relevant to the present force reconstruction with economical sampling density. The traction recovery for a single cardiac myocyte, which is in good agreement with that obtained by the Fourier transform traction cytometry, demonstrates the feasibility of the proposed method. We have developed a simple and efficient method to recover cellular traction field from substrate deformation. Unlike previous force reconstructions that numerically employ some regularization schemes, the present approach stabilizes the traction recovery by analytically improving the Green’s function such that the intricate regularizations can be avoided under proper conditions. The method has potential application to a real-time traction force microscopy in combination with a high-efficiency displacement acquisition technique.

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