Determination of oxidative stress in human lymphocytes exposed to Isocyanates

In present work it was found that during environmental stress, ROS levels can be increased, resulting in damage to cellular structures where Glutathione levels get diminished. The dye having high affinity for glutathione fluoresced Blue when bound to reduced or oxidized glutathione whereas unbound dye remained non-fluorescent. Thus, decrease in glutathione level could be detected in apoptotic cells. Similarly, measurement of 8-oxo-dG indicated oxidative damage where an 8-oxo-dG antibody bound to the 8-oxo- dG in the sample, standard/prebound to the wells. Anti-8-oxo-dG bound to 8-oxo-dG washed away while those captured by the immobilized 8-oxo-dG were detected by a secondary antibody: HRP conjugate. The intensity of yellow color was inversely proportional to the concentration of 8-oxo-dG. Another measure of oxidative stress is CM- H2DCFDA; a cell permeant, nonfluorescent indicator, which is redox sensitive dye until removal of the acetate groups by intracellular esterases, and oxidation occurred within the cell.