Specimen Preparation for Single-Cell Sequencing Analysis of Skeletal Cells

Recent work emphasizes that bone comprises numerous mesenchymal cell types each with different biologic functions, and deconvoluting the functions of these cells requires technical approaches with single-cell resolution, such as single-cell RNA sequencing (scRNA-seq). A critical step in conducting a successful single-cell sequencing study of bone is generation of a single-cell suspension of skeletal cells while preserving cell viability. Here we describe a method to prepare single-cell suspensions from skeletal tissue in preparation for single-cell sequencing studies. Also included are optional steps for fluorescence-activated cell sorting (FACS) of skeletal cells, which allows subsequent scRNA-seq studies to be focused on specific populations of interest.