Copyright q American Society of Andrology Utility of the Nitroblue Tetrazolium Reduction Test for Assessment of Reactive Oxygen Species Production by

ABSTRACT: The purpose of this study was to evaluate the ability of spermatozoa and leukocytes in semen to produce reactive oxygen species (ROS) by using nitroblue tetrazolium (NBT) staining and to examine the association between NBT staining and levels of ROS as measured by chemiluminescence. Twenty-one infertility patients (leukocytospermia; n 5 8; nonleukocytospermia, n 5 13) and 9 healthy donors were included. Standard semen analysis and density gradient centrifugation were performed to test NBT staining, ROS, and total antioxidant capacity. A ROS–total antioxidant capacity (ROS-TAC) score was calculated by using principal component analysis. In the leukocytospermic group, after separation on a den-sity gradient, the percentage of NBT-positive staining was signifi-cantly higher in sperm suspensions contaminated with leukocytes (median [25th, 75th percentiles]; 70 % [61%, 79%]) compared to the nonleukocytospermic group (14.5 % [9%, 25.5%]; P 5.03) and donor