Real‐time tracking of multiple particles is key for quantitative analysis of dynamic biophysical processes and materials science via time‐lapse microscopy image data, especially for single molecule biophysical techniques, such as magnetic tweezers and centrifugal force microscopy. However, real‐time multiple particle tracking with high resolution is limited by the current imaging processes or tracking algorithms. Here, we demonstrate 1 nm resolution in three dimensions in real‐time with a graphics‐processing unit (GPU) based on a compute unified device architecture (CUDA) parallel computing framework instead of only a central processing unit (CPU). We also explore the trade‐offs between processing speed and size of the utilized regions of interest and a maximum speedup of 137 is achieved with the GPU compared with the CPU. Moreover, we utilize this method with our recently self‐built centrifugal force microscope (CFM) in experiments that track multiple DNA‐tethered particles. Our approach paves the way for high‐throughput single molecule techniques with high resolution and efficiency.
Particles are widely used as probes in life sciences through their motions. In single molecule techniques such as optical tweezers and magnetic tweezers, microbeads are used to study intermolecular or intramolecular interactions via beads tracking. Also tracking multiple beads’ motions could study cell–cell or cell–ECM interactions in traction force microscopy. Therefore, particle tracking is of key important during these researches. However, parallel 3D multiple particle tracking in real‐time with high resolution is a challenge either due to the algorithm or the program. Here, we combine the performance of CPU and CUDA‐based GPU to make a hybrid implementation for particle tracking. In this way, a speedup of 137 is obtained compared the program before only with CPU without loss of accuracy. Moreover, we improve and build a new centrifugal force microscope for multiple single molecule force spectroscopy research in parallel. Then we employed our program into centrifugal force microscope for DNA stretching study. Our results not only demonstrate the application of this program in single molecule techniques, also indicate the capability of multiple single molecule study with centrifugal force microscopy.