Objective: The purpose of this study was to determine whether five-color flow cytometry could be used to examine the effect of acute Tai Chi (TC) practice on pro-inflammatory monocyte subtypes within the total monocyte population in peripheral blood. Design: A pilot pre- post-intervention design was our experimental model: pre-intervention measurements acted as the control for the post-intervention measurements. The research was conducted at Landsbaum Center for Health Education in a Midwestern community. Healthy subjects were asked to perform their style of TC for one hour. Methods: Four milliliter of peripheral blood was collected in the BD Vacutainer CPT tubes immediately prior and after one hour of TC practice. Peripheral blood mononuclear cells (PBMCs) were isolated following the manufacturer's protocol. PBMCs were then assessed for CD14, CD16, TLR2, TLR4, and HLA-DR expressions using five-color flow cytometry measurement. FlowJo and Graphpad Prism software were used to analyze the results. Results: The proportion of pro-inflammatory CD14+CD16+ monocytes decreased immediately after one hour of TC practice (p<0.01). Conclusion: The results indicate that the five-color flow cytometric process was sufficiently sensitive to be used to explore changes in peripheral blood pro-inflammatory monocytes following one hour of TC practice.