Semen from 4 Holstein bulls was diluted in 4 different extenders, filtered with Sephadex ion-exchange column, and frozen in liquid nitrogen. Sperm motility, progressive motility, path velocity, progressive velocity and the percentage of normal acrosomes of filtered and nonfiltered semen were recorded before and after freezing. Semen characteristics were significantly influenced by extender, filtration and freezing. Before and after freezing, motility measurements and the percentage of normal acrosomes were higher (P<0.001) in filtered than in nonfiltered spermatozoa. Post-thaw recovery rate of:motile spermatozoa was higher in filtered semen than nonfiltered (68 vs 39%, P<0.0001). The reduction in motility, progressive motility and the percentage of normal acrosomes during freezing and thawing processes were significantly lower (P<0.0001) in filtered semen (34, 34 and 4%, respectively) than nonfiltered (59, 54 and 15%, respectively). Post-thaw viability of spermatozoa was significantly affected by extender, filtration and time (P<0.0001). Immediate (0 h) post-thaw motility of nonfiltered semen (29%) was similar to 4-h post-thaw motility of filtered semen (25%; P>0.05). In conclusion, bull spermatozoa recovered by Sephadex ion-exchange filtration showed better post-thaw viability.