Offspring from Mouse Embryos Developed Using a Simple Incubator-Free Culture System with a Deoxidizing Agent

Abstract

To culture preimplantation embryos in vitro, water-jacketed CO2 incubators are used widely for maintaining an optimal culture environment in terms of gas phase, temperature and humidity. We investigated the possibility of mouse embryo culture in a plastic bag kept at 37°C. Zygotes derived from in vitro fertilization or collected from naturally mated B6D2F1 female mice were put in a drop of medium on a plastic culture dish and then placed in a commercially available plastic bag. When these were placed in an oven under air at 37°C for 96 h, the rate of blastocyst development and the cell numbers of embryos decreased. However, when the concentration of O2 was reduced to 5% using a deoxidizing agent and a small oxygen meter, most zygotes developed into blastocysts. These blastocysts were judged normal according to their cell number, Oct3/4 and Cdx2 gene expression levels, the apoptosis rate and the potential for full-term development after embryo transfer to pseudopregnant recipients. Furthermore, using this system, normal offspring were obtained simply by keeping the bag on a warming plate. This culture method was applied successfully to both hybrid and inbred strains. In addition, because the developing embryos could be observed through the transparent wall of the bag, it was possible to capture time-lapse images of live embryos until the blastocyst stage without needing an expensive microscope-based incubation chamber. These results suggest that mouse zygotes are more resilient to their environment than generally believed. This method might prove useful in economical culture systems or for the international shipment of embryos.

Oocyte collection and in vitro maturation after train transportation of human follicular fluid aspirated from resected non‐stimulated ovaries of patients with endometrial adenocarcinoma

“…Porcine follicular fluid that was kept at a low temperature for a long time was found to result in reduced oocyte quality . Also, it was reported that high pH in the transportation environment lowered embryo quality . In this study, we mixed HFF with maturation medium in order to prevent evaporation and to improve the HFF environment during transportation.…”

“…6 Also, it was reported that high pH in the transportation environment lowered embryo quality. 30 In this study, we mixed HFF with maturation medium in order to prevent evaporation and to improve the HFF environment during transportation. Additional research is required to determine whether HFF should be transported alone or mixed with maturation medium.…”

Decisions for the IVF laboratory: comparative analysis of embryo culture incubators

“…Incubator selection is an important decision for the IVF laboratory, as these devices regulate several environmental variables that can impact embryo development. While novel culture approaches may reduce the need for traditional incubators (Blockeel et al, 2009;Hyslop et al, 2012;Itoi et al, 2012;Ranoux and Seibel, 1990;Ranoux et al, 1988;Suzuki et al, 1999;Swain, 2010Swain, , 2011Taymor et al, 1992;Vajta et al, 1997Vajta et al, , 2004Van Blerkom et al, 2013;Varisanga et al, 2000), for the time being they remain a central part of a modern IVF laboratory. Functional aspects of the incubator, such as gas capability and sensor type, as well as temperature control and size/patient capacity need to be considered.…”

A Simple Method for Transportation of Mouse Embryos Using Microtubes and a Warm Box

“…On the day of embryo transfer, recipients were anesthetized by intraperitoneal injections of 0.02 mL/g of tribromoethanol. Five to 10 embryos were transferred into each uterine horn [ 4 , 13 ]. Offspring were obtained at 18.5 dpc via Cesarean section, and the body and placental weights were recorded.…”

“…Recently, we reported the use of a simple incubator-free culture system with a deoxidizing agent [ 4 ]. In this system, most mouse embryos developed into blastocysts in a plastic bag without the need for maintaining humidity, and healthy offspring were obtained.…”