The hypo-osmotic swelling test was claimed to assess an independent functional characteristic of human spermatozoa bearing relevance to their fertilizing capacity. To test this claim, we have studied the relationship between the result of the hypo-osmotic swelling test with that of conventional semen analysis and sperm motility patterns, the semen content of adenosine triphosphate, the staining pattern to acidified aniline blue, and the zona-free hamster oocyte test. The result of the HOS test is significantly correlated with all sperm characteristics except for the aniline blue stainability and the hamster oocyte test. The capacity of spermatozoa to react in a hypo-osmotic environment expresses the same functional information as the viability test using eosine staining. It is concluded that the hypo-osmotic swelling test does not add relevant information to that obtained by routine sperm analysis with regards to the fertilizing potential of semen.