Pm. Zavos et al., EPIDIDYMAL SPERMATOZOA - RECOVERY AND SUBSEQUENT IMPROVEMENTS OF MOUSE EPIDIDYMAL SPERMATOZOA VIA THE SPERMPREP(TM) FILTRATION METHOD, Tohoku Journal of Experimental Medicine, 175(2), 1995, pp. 101-109
This study was designed to determine the effects of Sephadex filtratio
n (SpermPrep(TM) method) on the separation of viable motile, morpholog
ically normal mouse epididymal spermatozoa and to study the viability
of the recovered spermatozoa over a 3 hr incubation period. Spermatozo
a were harvested from the caudae epididymides (5 animals per run or re
plication; n=10) following bilateral testicular excision and incubated
in 2-ml of Test-Yolk buffer (TYB) at 37 degrees C for 15 min. The spe
cimen was then split into 2 aliquots, with Alquot 1 as the control and
Aliquot 2 used for filtration. SpermPrep(TM)I column was employed acc
ording to the manufacturer's specifications (ZBL, Inc., Lexington, KY,
USA) using TYB. During filtration (10 min), different fractions were
obtained: first 5 min (Sample 1) and second 5 min (Sample 2). The filt
ered fractions were evaluated and incubated at 37 degrees C and assess
ed for percentage and grade of motility (0-4) every 30 min for 3 hr. F
iltration resulted in a significant improvement in percentage and grad
e of motility (91.5% and 3.0 vs. 76.5% and 2.5, respectively). The res
ults point out very clearly that the filtration via the SpermPrep(TM)
method improved the percentage and grade of motility (p<0.05) but not
the percentage normal morphology of the spermatozoa. Also the SpermPre
p(TM)I enabled the recovery of 45% (8.3 x 10(6) spermatozoa) of the to
tal spermatozoa processed in the control aliquot (18.4 x 10(6) spermat
ozoa) which is consistent with previous observations. Most importantly
, filtered spermatozoa incubated for 3 hr showed greater percentage an
d grade of motility than the control spermatozoa (63% and 1.66 vs. 39%
and 0.82, respectively). This is of significant importance since the
filtration via the SpermPrep(TM)I has selected not only greater qualit
y spermatozoa but also spermatozoa with significantly long-term motili
ty (longevity) during in vitro incubation.