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The MTT assay application to measure the viability of spermatozoa: A variety of the assay protocols

Kakanang Buranaamnuay


The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay is one of the methods used to evaluate the viability of sperm. In the assay, a tetrazolium component (MTT) is converted into MTT formazan by some specific enzymes in the viable cells. The amount of formazan product in theory is directly correlated with the percentage of viable sperms. It is quantified by measuring the absorbance using a spectrophotometer. The present article compiles the MTT assays that have been used to determine sperm viability in most animal species and humans. In each assay, other factors apart from the number of viable cells that potentially influence the accuracy and precision of results are stated, such as preparations of sperm and MTT solutions, length and conditions of incubation, and a solubilizing agent as well as the formazan detection method. Also, the strengths and shortcomings of the MTT test comparison with the others are summarized at the end of this article. This information may be useful for prospective researchers deciding to implement this colorimetric method in their experiments.