The use of iodixanol as a density gradient material for separating human sperm from semen

Iodixanol, a new nonionic density gradient material with relatively low osmolality and high density, was evaluated to determine its suitability for the separation of human sperm from semen for their subsequent therapeutic use. Using a three-layer iodixanol gradient (approximately 1.17/1.15/1.05 g/mL), sperm were centrifuged at 1000g for 30 min and collected from the 1.05/1.15 interface. Using this method, a mean of 78% of the motile and 99% of the morphologically normal sperm originally present in the semen were recovered at the interface. There was no significant increase in the percentage of motile or morphologically normal sperm in the final preparation compared to the original semen. Sperm survived iodixanol density gradient centrifugation well, showing only modest declines in motility (18%) and velocity (35%) during a subsequent 24-h incubation period. When iodixanol was compared to Percoll density gradient centrifugation with semen from the same ejaculate, there was no significant difference between methods with regard to sperm yield, enrichment of motile or morphologically normal sperm in the preparation or sperm survival following separation. Iodixanol provides a suitable, nontoxic alternative to Percoll for the preparation of human sperm for therapeutic use.