Suitability of the hypo-osmotic swelling test for assessing the viability of cryopreserved sperm.
OBJECTIVE
To determine whether the hypo-osmotic swelling test can assess viability of frozen spermatozoa.


METHODS
Fresh semen specimens were divided into two aliquots: one was not processed and the other was processed by swim-up. Both aliquots were then cryopreserved by the liquid nitrogen vapor method.


METHODS
Andrology laboratory at a tertiary care facility.


METHODS
Eleven volunteers with proven fertility.


METHODS
The hypo-osmotic swelling test and the sperm viability assessment by Hoechst 33258 (Sigma Chemical Co., St. Louis, MO) and eosin-nigrosin strains were performed in each aliquot.


METHODS
The results of hypo-osmotic swelling test as indicated by swelling of sperm tails were compared with the viability results obtained by Hoechst 33258 stain before freezing and by Hoechst-33258 and eosin-nigrosin stains after freezing.


RESULTS
The hypo-osmotic swelling test and Hoechst 33258 viability results were correlated highly in unprocessed and processed fresh specimens (r = 0.95). After cryopreservation, there was no correlation between hypo-osmotic swelling test and Hoechst 33258 or between hypo-osmotic swelling test and eosin-nigrosin. Prefreeze sperm processing did not influence the post-thaw results. Viability assessed by Hoechst 33258 and eosin-nigrosin was well correlated (r = 0.72).


CONCLUSIONS
The hypo-osmotic swelling test can accurately evaluate viability in fresh human spermatozoa but not in cryopreserved ones. Thus, this test cannot be used to select viable cells in cryopreserved samples for assisted reproductive procedures, such as intracytoplasmic sperm injection.
Esteves
S C
SC
Department of Urology, Cleveland Clinic Foundation, Ohio 44195, USA.
Sharma
R K
RK
Thomas
A J
AJ
Agarwal
A
A
D000814:Aniline Compounds; D006690:Bisbenzimidazole; D002470:Cell Survival; D004396:Coloring Agents; D015925:Cryopreservation; D004801:Eosine Yellowish-(YS); D006801:Humans; D007038:Hypotonic Solutions; D008297:Male; D013082:Sperm Tail; D013094:Spermatozoa / Q000502:physiology*
D000814:Aniline Compounds; D004396:Coloring Agents; D007038:Hypotonic Solutions; C002712:nigrosin; D006690:Bisbenzimidazole; D004801:Eosine Yellowish-(YS)