Fig. 1

Experimental study design. Samples were obtained from 33 men undergoing routine semen analysis. Excess semen from each sample was split in two and processed in parallel in two ways: a control arm “H -”: sperm cells were processed by density gradient centrifugation (DGC) without hypotaurine supplementation, and an “H +” arm: spermatozoa were processed by 50 mM hypotaurine supplementation during DGC. Spermatozoa were washed and frozen in high-security straws in the presence of hypotaurine supplemented (H+) or non-hypotaurine supplemented (H-) media. After thawing, the different analyzes were carried out in parallel for both conditions: - (i) Analysis of standard semen parameters (n = 33): Sperm vitality,progressive and total motilities. - (ii) Measurements of fertilization capacity: Evaluation of (cryo-) capacitation terminal markers by measuring tyrosine phosphorylation of P110 and P80 (n = 15) and analysisof acrosome integrity after FITC- PSA labeling (n = 25), and - (iii) Measurements of nuclear quality markers: Chromatin condensation measured by chromomycin A3 (n = 28), DNA fragmentation by TUNEL assay (n = 18), and nuclear vacuolization observed by examination of motile sperm organelle morphology (MSOME, n = 19)