Development of a cellular force measurement system through elastic micropillars and nanodiamond nitrogen-vacancy centers

Abstract

In order execute biology duties such as migration and proliferation, cells need to establish stable adhesion with outside and then exert contractile forces (typically in the pico to nano newton range) to probe their micro-environment. However, how to accurately measure such forces, especially the component normal to the cell-substrate interface, remains challenging. In recent years, the concept of utilizing fluorescent nanodiamond nitrogen-vacancy (NV) centers in tomographic vector magnetometry to monitor nano-scale three-degrees-of-freedom (3-DoF) motion and deformation has received increasing attention. Physically, the rotation of an object can be tracked by a set of Euler, namely, the roll, pitch, and yaw angles. Since the orientation of a NV center (representing an impurity in diamond where 2 carbon atoms are replaced by a nitrogen-vacancy) can be detected by its luminescent signals under excitation, such structure becomes ideal for us to accurately measure the aforementioned Euler angle changes at nano-scale. In this study, we designed and fabricated a cellular force measurement tool that combines NV centers with PDMS micropillar arrays. The essential idea is to attach nanodiamonds (NDs) containing NV centers to the top of PDMS micropillars, so that the position and angle change of each pillar can be detected via magnetic resonance (ODMR) measurement, using NV centers as markers, which eventually allows us to precisely estimate the force acting on the pillar. To achieve these goals, several methods for combining NDs with micropillars were developed and tested. The first is to mix NDs with PDMS before curing which results in the aggregation of NDs, due to gravity, at the bottom of silicon mould and therefore in the top of pillars after demoulding. The second method is to anchor NDs on the top surface of pillars through covalent bonding between BCN and azide on a surface modification polymer[206]. The third one is to use dopamine hydrochloride to connect NDs and pillars by self-aggregation. We then summarize and demonstrate the advantages and disadvantages of each method. The successful fabrication of NDs-coated pillars was confirmed by fluorescence imaging of NV centers. In addition, the bio-compatibility of the system was demonstrated by growing NIH 3T3 cells on the pillar array. Interestingly, scanning electron microscope (SEM) examination showed that cells could spread well on the top of multiple pillars, indicating the suitability of our system in achieving cellular force measurement eventually. Finally, we have also preliminarily tried to connect the force acting on the pillar with its magnetic signal change associated with NV centers. Specifically, atomic force microscopy (AFM) was used to push and deform specifically fabricated PDMS micropillars on a coverslip. Since the force exerted by AFM can be precisely controlled, this will allow us to calibrate and validate the cellular force measurement capability of the system.