Optimization of Cytometry by Time-of-Flight Staining for Peripheral Blood and Bone Marrow Samples.

Cytometry by time-of-flight (CyTOF) enables comprehensive immune profiling for translational research. However, challenges such as signal variability, nonspecific binding, and antibody incompatibility can compromise data quality. This study presents an optimized CyTOF staining protocol for human peripheral blood mononuclear cells and bone marrow aspiration concentrate samples, addressing these challenges by refining antibody conjugation with polymer X8, saponin use, and fixation protocols. Preliminary data indicate improved staining for key markers (CD14, CD16, and CD19), enhancing signal consistency and clarity. These findings advance the utility of CyTOF in orthopaedic research and immune profiling for diseases such as osteonecrosis of the femoral head.