Comparison of four fluorochromes for the detection of the inner mitochondrial membrane potential in human spermatozoa and their correlation with sperm motility

BACKGROUND: Sperm motility evaluation is associated with fertility in IVF programmes. The visual estimation of sperm motility is extremely subjective. Hence, alternative methods are required. Among them, determination of mitochondrial membrane potential (Dcm) changes of spermatozoa using potentiometric dyes may be a reliable test to determine sperm quality. However, the use of the potentiometric dyes in sperm samples has not been compared. METHODS: We have studied sperm samples from 28 infertile patients enrolled in an IVF programme in flow cytometry after staining of spermatozoa with four commonly used potentiometric dyes. Sperm motility was evaluated visually. RESULTS: As expected, JC-1 seems to detect specifically Dcm changes, CMX-Ros, DiOC6(3) and TMRE fluorescence is easily analysed and the latter three fluorochromes are particularly suitable for multiparametric staining. Irrespective of the Dcm-dependent fluorochromes used to stain spermatozoa, a positive correlation was found between the percentage of Dcm high cells and forward motility and also with high fertilization rates after IVF. CONCLUSION: The four fluorochromes may be useful for evaluation of sperm samples from infertile patients. The choice of the potentiometric dyes will depend on their fluorescence characteristics in order to use them in combination with other fluorescent markers.