QUANTIFICATION OF REACTIVE OXYGEN SPECIES GENERATED BY ALVEOLAR MACROPHAGES USING LUCIGENIN-ENHANCED CHEMILUMINESCENCE - METHODICAL ASPECTS

Alveolar macrophages lavaged from bovine lungs using Ca2+- and Mg2+-de prived saline containing EGTA for calcium chelation were cultivated in RPMI-1640 medium. The generation of reactive oxygen species (ROS) was assessed by determination of lucigenin-enhanced chemiluminescence (LU C-CL) using a LB 9505 C Biolumat. Monitoring of LUC-CL depends on vari ous methodical parameters: besides medium constituents, the method of cell harvesting and the time-schedule of the protocol appear to be ess ential parameters that influence ROS-generation. In addition, the cell number (cell density) as well as the ratio of cell number to particle mass influence the amount of ROS-generation. Following exposure of ce lls to micronized quartz, we observed a dose-related increase in the g eneration of ROS.