Human Reproduction, Vol 12, 1550-1553, Copyright © 1997 by Oxford University Press
PL Matson, J Graefling, SM Junk, JL Yovich and WR Edirisinghe
Mouse oocytes and embryos were obtained following ovulation induction of
(C57B16 x CBA) F1 animals. Zonae pellucidae were exposed to alpha-
chymotrypsin in phosphate-buffered medium (PB1) supplemented with 3 mg/ml
bovine serum albumin upon a heated stage (37 degrees C) and were observed
constantly through an inverted microscope. The endpoint of the bioassay was
the limits of the zona no longer being seen clearly at x 200 magnification,
and the time taken for each zona to dissolve was recorded. A dose-dependent
response in dissolution time was clearly seen, with 1% alpha-chymotrypsin
being chosen as the routine working solution. Cryopreservation of 2-cell
mouse embryos using propanediol did not cause zona hardening but induced a
small and significant softening, as gauged by the time taken for zona
dissolution (2181 +/- 167 versus 1864 +/- 82 s). Zona hardening was not
suspected to occur after the freezing of human embryos as there was no
difference in implantation rates per embryo for in-vitro fertilization
(IVF) and intracytoplasmic sperm injection (ICSI) treatment cycles between
fresh [IVF: 63/644 (9.7%); ICSI: 51/330 (15.5%)] and frozen embryos [IVF:
36/458 (7.9%); ICSI: 18/112 (16.1%)]. Conversely, significant hardening of
the zonae of mature oocytes was seen following cryopreservation (747 +/-
393 s) compared with freshly ovulated oocytes (151 +/- 68 s). It is
concluded that (i) the freezing of murine oocytes with propanediol results
in zona hardening, implying a possible benefit of ICSI after the
cryopreservation of human oocytes, and (ii) the cryopreservation of embryos
is not associated with zona hardening or reduced implantation, making
microdissection of the zona in such cases generally unwarranted.
ARTICLES
Cryopreservation of oocytes and embryos: use of a mouse model to investigate effects upon zona hardness and formulate treatment strategies in an in-vitro fertilization programme
PIVET Laboratory, Leederville, Western Australia.
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