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Animal Reproduction Science 60–61 2000 81–91 www.elsevier.comrlocateranireprosci
In vitro culture of bovine embryos in Menezo’s B2
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medium with or without coculture and serum: the normalcy of pregnancies and calves resulting from
transferred embryos
John F. Hasler
Em Tran, Inc., 197 Bossler Road, Elizabethtown, PA 17022, USA
Abstract
The present study was designed to test the efficacy, as assessed by blastocyst production, of Ž
. culturing slaughterhouse-derived bovine oocytes in Menezo’s B2 B2 medium with or without
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serum and with or without buffalo rat cell BRL coculture. In addition, OPU-derived oocytes were cultured in B2–BRL coculture with or without serum for the first 72 h or TCM 199–BRL
coculture with serum and the resulting embryos were transferred into recipients. Culture in B2 plus serum resulted in more blastocysts than B2 without serum, but both treatments produced far
fewer blastocysts than did B2–BRL coculture. In addition, B2 with or without serum produced embryos of a lower stage of development, lower quality and fewer cells than did B2–BRL
coculture. Serum restriction for the first 72 h of B2–BRL coculture did not result in a detectable difference in blastocyst production during 7 or 8 days of culture relative to coculture with no
serum restriction. Embryos produced in B2–BRL coculture with serum restriction resulted in pregnancy rates, percentage of male fetuses, abortions, and congenital problems similar to those
from B2 or TCM 199–BRL coculture with no serum restriction. q 2000 Elsevier Science B.V. All rights reserved.
Keywords: Cattle; In vitro; Embryo culture; Pregnancies; Calves
1. Introduction
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In vitro culture IVC techniques for the production of cattle embryos became widely used on a commercial basis in a number of countries during the 1990s. Although
Fax: q1-717-653-6554. Ž
. E-mail address: jfhaslerredrose.net J.F. Hasler .
0378-4320r00r - see front matter q 2000 Elsevier Science B.V. All rights reserved. Ž
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thousands of calves have been produced, problems with abortions, increased birth Ž
weights, dystocias and higher rates of neonatal mortality have been widely reported for .
reviews, see Walker et al., 1996; Farin and Farin, 1997; Kruip and den Daas, 1997 . Ž
. Walker et al. 1992 first described increased birth weights, lengthened gestation periods
and higher neonatal mortality in sheep pregnancies resulting from in-vitro-derived pregnancies. Subsequently, the inclusion of serum during the in vitro culture of ovine
Ž embryos was implicated in these abnormalities Thompson et al., 1995; Holm et al.,
. 1996 . As a result, there have been a number of recent studies directed at more clearly
Ž . defining the problem s of the inclusion of serum in the in vitro culture of both ovine
Ž and bovine embryos Popovic et al., 1997; Thompson et al., 1998; Blondin et al., 1999;
Jacobsen et al., 1999; Sinclair et al., 1999; Tricoire et al., 1999; van Wagtendonk-de .
Leeuw et al., 1999 . It remains to be demonstrated that any of the currently used in vitro systems result in the production of a statistically significant improvement in the number
of ‘normal’ pregnancies. The present study was organized to determine the efficacy of the in vitro culture of bovine zygotes with and without somatic cell coculture and with
or without serum for the first 72 h of culture. In addition, a survey was conducted on whether the restriction of serum during the first 72 h of development in Menezo’s B2
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B2 –buffalo rat cell BRL coculture improved the normalcy of pregnancies resulting from the transfer of embryos.
2. Materials and methods
