ON TESTIS AND EPlDlDYMlS OF RATS - Pondicherry University ...

choppcd with the help of sharp razor blude in 5 mL ol' Ilam's 1:12 medium and
incubated for 5 min at 35'C. After several washings in the Ham's F12 medium.
sperm collected in the medium were used for various experiments.
3.11.1 Sperm viability test
Sperm viability test was done by the method as described in the WHO
Laboratory Manual (1999). An aliquot of 100 pL of epididymal sperm was mixed
with I00 pL of 0.5% eosin solution. A drop ofthe ubovc mixturc wils put on a n1icl.oslide
covered with a cover slip and examined after 30 sec at 200 x with a help of light
microscope. Randomly two hundred spermatozoa were counted. The livc
spermatozoa were unstained and dead cells were stained. The sperm viability was
expressed in percentage as the number of viable sperm of the total sperm counted.
3.11.2 Epididymal sperm motility
Motility of epididymal sperm was evaluated by the method as described by
Linder et a/. (1986) and Cooke el al. (1991). A small incision was made in the cauda
epididymis using a sharp razor. The fluid, which oozed out from the cauda
epididymis was taken by using a pipene tip and diiuted to 2 mL with Ham's F12
medium at 3S°C. An aliquot of this solution was placed in a hemocytometer and
counted for motile and non-motile sperm. First non-motile sperm were counted
followed by motile sperm. Epididymal sperm motility was expressed as the
percentage of motile sperm of the total sperm counted.
3.1 1.3 Epididymal sperm counts
Epididymal sperm were counted by the method as described in the WHO
Laboratory Manual (1999). An aliquot of 5 111. of epididyn~al sperm was diluted with
95 pL of diluents (see Appendix 1.2). A coverslip was placed on the counting
chambers of the improved Neubauer type hemocytometer.