Abstract
Chemiluminescence (CL) in whole blood has been analyzed to assess the phagocytic function of granulocytes and the serum opsonic activity simultaneously. Granulocytes are well known to show CL during phagocytosis, which is derived from the interaction between activated oxygen species generated in cells and substances phagocytosed. The total activity of activated oxygen species generated from phagocytes can be measured in luminol-dependent CL. This method, namely luminol-dependent CL, has become an elegant and simple method to estimate the phagocytic function of granulocytes or monocytes. The method described here is an improved one, in which whole blood can be directly used and serum opsonic activity can be measured simultaneously. The methods are as follows; a 0.1ml of freshly drawn whole blood is put into a CL measuring vial, in which a 0.4ml of medium (Eagle's medium buffered with 50mM HEPES containing 10U/ml of heparin) has been put. After 20min incubation at 37°C, 10μl of luminol at the final concentration of 40μg/ml is added to a specimen. Following equilibration for 10min, a background CL is measured. Then a 10μl of stimulant is added to the specimen. CL is continuously recorded by a recorder connected to the CL machine (BLR-102, Aloka). The results obtained in this study are as follows;
1) The peak CL in whole blood is about 1/30 of that of granulocytes alone, however, the CPM is high enough for the estimation of phagocytic function of granulocytes.
2) The peak CPM correlates to the number of granulocytes present in specimen, and/or function of granulocytes.
3) The time showing the peak CL depends on the activity of serum opsonin.
4) If aggregated gamma globulin is used as the stimulant, it can quickly activate granulocytes without opsonin.
5) Sheep red blood cells can be a stimulant, if complement and specific antibody are present in blood. These results seem to confirm that the measurement of CL in whole blood offers much information concerning functions of phagocytic system in whole blood.
