Deep Learning Image Recognition-Assisted Atomic Force Microscopy for Single-Cell Efficient Mechanics in Co-culture...

Atomic force microscopy (AFM)-based force spectroscopy assay has become an important method for characterizing the mechanical properties of single living cells under aqueous conditions, but a disadvantage is its reliance on manual operation and experience as well as the resulting low throughput. Particularly, providing a capacity to accurately identify the type of the cell grown in co-culture environments without the need of fluorescent labeling will further facilitate the applications of AFM in life sciences. Here, we present a study of deep learning image recognition-assisted AFM, which not only enables fluorescence-independent recognition of the identity of single co-cultured cells but also allows efficient downstream AFM force measurements of the identified cells. With the use of the deep learning-based image recognition model, the viability and type of individual cells grown in co-culture environments were identified directly from the optical bright-field images, which were confirmed by the following cell growth and fluorescent labeling results. Based on the image recognition results, the positional relationship between the AFM probe and the targeted cell was automatically determined, allowing the precise movement of the AFM probe to the target cell to perform force measurements. The experimental results show that the presented method was applicable not only to the conventional (microsphere-modified) AFM probe used in AFM indentation assay for measuring the Young's modulus of single co-cultured cells but also to the single-cell probe used in AFM-based single-cell force spectroscopy (SCFS) assay for measuring the adhesion forces of single co-cultured cells. The study illustrates deep learning imaging recognition-assisted AFM as a promising approach for label-free and high-throughput detection of single-cell mechanics under co-culture conditions, which will facilitate unraveling the mechanical cues involved in cell-cell interactions in their native states at the single-cell level and will benefit the field of mechanobiology.