Highly sensitive determination of plasma cytokines by time-resolved fluoroimmunoassay; Effect of bicycle exercise on plasma level of interleukin-1 alpha (IL-1 alpha), tumor necrosis factor alpha (TNF alpha), and interferon gamma (IFN gamma)

A highly sensitive time-resolved fluoroimmunoassay of human plasma cytokine s is described. The cytokines such as interleukin-1 alpha (IL-1 alpha), tum or necrosis factor alpha (TNF alpha) are known to be acute inflammatory cyt okines and it has been reported that these cytokines are secreted into bloo d by physical exercise. In this study, a sandwich-type immunoassay of cytok ines was established using a europium chelate BHHCT-Eu3+ as a powerful labe ling material. The minimum detection limits of cytokines, i.e. IL-1 alpha, TNF alpha, and interferon gamma (IFN gamma) were about 1/10 smaller than th ose of enzyme-linked immunosorbent assay currently used. By this immunoassa y we investigated cytokine increase/decrease in plasma which was thought to derive from the myocytes damaged by bicycle exercise. Healthy young men pe rformed two kinds of bicycle ergometer exercises, under conditions of an in cremental and a constant loading. Blood samples were taken before, during, and after exercises, and the concentration levels of plasma IL-1 alpha TNF alpha, and IFN gamma were determined. In the case of incremental exercise, IL-1 alpha increased significantly at the first stage but decreased to the basal level from the second stage, in spite of heavier exercise. In the cas e of 30 min constant exercise, the level of plasma IFN gamma increased in r ecovery period, 2 h after the light-exercise. TNF alpha level was significa ntly higher in a heavy-exercise. The concentration of IL-1 alpha peaked at the early stage of the incremental exercise; this fact has not been reporte d in previous studies. This cytokine is unique in showing a sudden increase during the early stage, while others increase after the exercise. Our high ly sensitive assay made it possible to detect a slight change in plasma cyt okines.