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A origem dos Complexos Cumulus-Oocitos (CCO) bovinos e os diferentes protocolos entre equipes geram variacoes nos indices de producao in vitro (PIV). O processo pode ser influenciado pelo numero de CCO e volume das gotas de maturacao, fecundacao e cultivo. Neste estudo, avaliou-se a densidade de CCO e o volume de meio de fecundacao sobre a PIV com 672 oocitos bovinos. A maturacao in vitro (MIV) foi conduzida com 20 CCO em 200µL de TCM-199+rFSHh+10% de soro de vaca em estro (SVE), por 24h, em estufa a 39oC, 5% CO2 em ar e umidade saturada. Na fecundacao (Dia 0 =D0) utilizou-se 5, 10 ou 20 CCO e 2 volumes de meio (1:5 e 1:10), num fatorial 3x2, em 6 repeticoes. O semen foi selecionado por migracao ascendente e para a fertilizacao (1x106 espermatozoides/mL) os gametas foram incubados em Fert-Talp, por 18h. O cultivo foi em grupos de 20 zigotos, em 200µL de fluido sintetico do oviduto (SOF) aaci+5%SVE, por 8 dias e as avaliacoes em D2 (clivagem), D7 (Blastocistos), D9 (Blastocistos expandidos, em eclosao e eclodidos). As taxas de clivagem (80, 85 e 87%), embrioes em D9 (23, 27 e 28%) e de eclosao (42, 37 e 48%) foram semelhantes (P>0,05) para 5, 10 ou 20 CCO, respectivamente. Nao houve diferenca (P>0,05) na clivagem (83 e 85%), embrioes em D9 (24 e 28%) e eclosao (41 e 44%) entre as proporcoes 1:5 e 1:10µL. A producao de blastocistos e taxa de eclosao em D9 nao sao influenciados pelo numero de CCO, nem pela proporcao de CCO por volume de meio na fecundacao. Volume of medium and number of oocytes for fertilization in the in vitro production of bovine embryos Abstract The origin of the bovine Cumulus Oocytes Complexes (COC) and the different protocols among teems are responsible for the variation rates of the in vitro production (IVP). The process is also influenced by COC number and by the volume of the drop in the maturation, fertilization and culture. In this study, the effect of bovine COC density per drop medium and the fertilization medium volume were evaluated during the bovine IVP using 672 COC. The in vitro maturation was performed with 20 COC in 200µL of TCM-199+rFSHh+10% estrus cow serum (ECS), for 24h in an incubator with 5%CO2 in air, saturated humidity, at 39oC. For the fertilization (Day 0 = D0) 5, 10 or 20 COC and 2 volumes ratio of medium (1:5 and 1:10) were used in a factorial outline 3x2, in 6 replications. Thawed semen was prepared by a swim up process and for the fertilization (1x106 spermatozoa/ ml) the gametes were maintained in Fert-Talp for 18h. Groups of twenty presumed zygotes were cultured in 200µL synthetic oviduct fluid (SOFaaci)+5% ECS, for 8 days. The evaluations were carried out in D2 (cleavage), D7 (blastocysts) and D9 (expanded and hatched blastocyts). The rates of cleavage (80, 85 and 87%), embryos in D9/ D2 (23, 27 and 28%) and hatched blastocstys in D9/D7 (42, 37 and 48%) were similar (P>0.05) for 5, 10 and 20 COC, respectively. No difference was observed in cleavage (83 and 85%), embryos in D9 (24 and 28%) and hatching rates (41 and 44%) for 1:5 and 1:10µL of fertilization medium. Blastocysts production and the hatching rate in D9 were neither influenced by the COC number, nor by the proportion of COC per medium volume used during fertilization.

VOLUME DE MEIO E NÚMERO DE OÓCITOS PARA FECUNDACÃO NA PRODUÇÃO IN VITRO DE EMBRIÕES BOVINOS