The influence of sperm/zona pellucida collision rates on zona binding
The satisfying success rates reported with intra-cytoplasmic sperm injection (ICSI) urged clinicians and scientists to re-address the emphasis in the management of the male factor patient towards gamete manipulation in order to circumvent the underlying problem causing fertilization failure. We have designed a study to (i) calculate the collision rate of a sperm population with the human zona pellucida, using a mathematical hypothesis and (ii) use the calculated collision rate to evaluate subsequent zona binding results obtained under hemizona assay conditions. Microdroplets were used to co-incubate sperm and human oocytes in order to evaluate zona binding. Using microvolumes, the track followed by sperm, as well as the maximum distance travelled were employed to calculate the collision rate of sperm and zona pellucida. The sperm concentrations of swim up samples were adjusted to 4 x 106 and 0.8 x 106 sperm ml-1. Five separate droplets each of 20 μl containing 4 x 106 sperm ml (80 000 motile sperm) and 200 μl containing 0.8 x 106 sperm ml-1 (160 000 motile sperm), respectively, were prepared. Both volumes were incubated for 18 h at 37 °C. The mean (± SD) number of spermatozoa tightly bound to hemizona, incubated in 20 μl and 200 μl sperm droplets, was 2444±612 and 548±315, respectively (P=0.0001). The results can be used as a guideline to calculate the optimum insemination concentration needed for a specific sperm population to ensure the maximum collision rates with the oocyte.