Purpose: No systematic investigation has been reported assessing the effect of cell isolation processes on postexercise apoptosis. Therefore, the effect of cell isolation procedures on apoptosis was evaluated in this study.
Methods: Untrained healthy individuals participated (N=13). Blood samples obtained at rest and immediately after an incremental exercise test to exhaustion were partitioned into three treatments: 1) whole blood smears made immediately after the sample was obtained (WB), 2) cells subjected to density-gradient isolation before smears were made (ISO), and 3) samples allowed to sit at room temperature (i.e., time-treated) before centrifugation and smearing (TT). Blood smears were stained using the May-Grünwald Giemsa procedure and lymphocytes were evaluated under a light microscope for characteristic features of apoptosis. Data were analyzed using a 2x3 ANOVA.
Results: A significant interaction effect existed (P<0.0001) such that at rest, no difference was detected in the amount of lymphocyte apoptosis among WB, ISO, or TT samples. However, after exhaustive exercise, the amount of apoptotic lymphocytes was significantly greater in WB compared with ISO and TT samples (P<0.0001).
Conclusion: Lymphocyte isolation results in a significant decrease in the percent of apoptotic lymphocytes after exhaustive exercise. This reduction is likely due to the time needed to isolate cells, rather than the isolation process itself. Because apoptosis is a time-sensitive process that occurs within minutes rather than hours, the length of time from initial sampling to the preparation of cells for assessment of apoptosis is critical and should be considered in future exercise studies.