A rapid microassay for measuring the luminol-dependent chemiluminescent response in canine whole blood

A microassay for the luminol-dependent chemiluminescence (CL) response in canine whole blood was developed to measure indirectly the oxidative metabolism of peripheral blood leukocytes. Fifty microliters of blood were mixed with 705 microliters of Hank's balanced salt solution containing 25 mM Hepes and 1.3 x 10(-4) M luminol. This mixture was allowed to equilibrate for 5 min after which 60 microliters of latex beads (0.801 microns diameter) were added as a stimulant, and the CL response was monitored continuously for 5 min at 37 degrees C using a luminometer. The whole blood CL response was significantly correlated (r = 0.784, P less than 0.01, n = 14) with the number of neutrophils in the peripheral blood. Further, the whole blood CL response was abolished by the depletion of neutrophils after passing the blood through an adherence column and by the addition of sodium azide. The relative chemiluminescent light unit (RCLU) was a reliable marker for comparing each peak value in different samples. The coefficient of variation (CV) of repetitive samples was 9.87%, and the CV of 14 normal dogs was 15.7%. This method is useful and applicable for screening the CL response in canine whole blood.