Sperm viability: a comparison of analytical methods

Two methods of measuring human sperm viability, the stain exclusion assay and the hypoosmotic swelling (HOS) test, were evaluated. Human sperm were pretreated with 2.0% glutaraldehyde or 0.1% Triton X-100 and compared to untreated controls. Approximately one half of the sperm were found to be viable in the control samples and nearly all sperm were non-viable in the Triton X-100 treated samples by both the stain exclusion and HOS assays. After glutaraldehyde pretreatment, presumably inactivating the spermatozoa, the HOS test revealed that most sperm were not viable, while the stain exclusion test found no difference between glutaraldehyde pretreated sperm and control sperm. Investigations with scanning and transmission electron microscopy demonstrated that the HOS test caused the membrane of the sperm tail to swell and the tail fibers to coil several times within the swollen membrane. It is concluded that the stain exclusion assay merely measures the structural integrity of the sperm membrane, whereas the HOS test also provides an indication of the physiological integrity of the sperm membrane.