Evaluation of chemiluminescence and flow cytometry as tools in assessing production of hydrogen peroxide and superoxide anion in human spermatozoa

Fertility and Sterility
Reda MahfouzAshok Agarwal

Abstract

To examine simultaneously the levels of hydrogen peroxide (H(2)O(2)) and superoxide (O(2)(-*)) using chemiluminescence and flow cytometry. Prospective laboratory study. Reproductive research lab in a tertiary hospital. Semen samples from 18 healthy male volunteers. Sperm preparation and measurement of reactive oxygen species (ROS) by chemiluminescence using luminol and lucigenin before and after H(2)O(2) exposure and by flow cytometry using dichlorofluorescin diacetate (DCFH-DA) for H(2)O(2) and dihydroethidium (DHE) for O(2)(-*). Sperm count, motility, viability, and ROS levels. Immature sperm fractions showed significantly higher levels of ROS measured by either luminol or lucigenin compared with the neat and mature fraction. ROS levels were detectable by flow cytometry in chemiluminescence-negative samples. Both mature and immature sperm fractions had a significantly higher percentage of cells positive for H(2)O(2) compared with neat semen. On the other hand, the percentage of O(2)(-*)-positive cells in neat semen was significantly higher compared with the percentage found in mature fractions but significantly lower than that in the immature sperm fractions. We recommend ROS measurement by flow cytometry on the basis that it...Continue Reading

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Citations

Dec 18, 2008·Applied Biochemistry and Biotechnology·Kavindra Kumar Kesari, J Behari
Mar 25, 2014·Colloids and Surfaces. B, Biointerfaces·Meghana RamaniEnrico Marsili
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