Enhanced chemiluminescence assay vs colorimetric assay for measurement of the total antioxidant capacity of human seminal plasma

Abstract

Although the enhanced chemiluminescence assay is commonly used to measure the nonenzymatic total antioxidant capacity (TAC) of the human seminal plasma, it is cumbersome, expensive, and time-consuming. We describe herein an alternate method to measure TAC that is based on the ability of antioxidants in seminal plasma to interfere with a reaction between 2,2'-azino-di-[3-ethylbenzthiazoline sulphonate] and metmyoglobin with H(2)O(2). This reaction produces a relatively stable blue-green color with absorbance maxima at 600 nm. We compared this colorimetric assay with our established chemiluminescence method and assessed quality control parameters (ie, intra-assay and interassay variabilities) in addition to intraobserver and interobserver differences. Our results show that the colorimetric assay was fairly predictive of antioxidant capacity similar to the chemiluminescence assay (P <.001). Furthermore, there was a high level of agreement between the duplicate measures by the same observer (intraobserver) and intra-assay variability, with a concordance correlation coefficient of 0.99. The interassay coefficient of variation was 4.7% (overall). The mean +/- SD of the difference between the 2 observers was 2.98% +/- 4.1%. In conclusion, we found that the colorimetric assay is a reliable and accurate method to evaluate seminal TAC, and it could be used as a simpler, rapid, and cheaper alternative to the chemiluminescence assay.