New pH-buffering system for media utilized during gamete and embryo manipulations for assisted reproduction

Maintenance of stable pH is important for optimizing gamete and embryo culture. One method to stabilize pH entails using zwitterionic buffers in IVF handling media used outside the laboratory incubator. Current handling media utilize single buffers, such as MOPS or HEPES. However, the use of a single buffer limits the ability to adjust the range of buffering capacity. Furthermore, changes in temperature alter buffering of these compounds. Therefore, traditional IVF handling media utilizing a single buffer may not provide ideal pH buffering. This study reports that combining multiple buffers, such as HEPES, MOPS and DIPSO, into a single medium in various ratios gives the ability to shift the effective buffering range to cover a specific pH. Additionally, by combining various buffers, it is possible to expand pH buffering over a range of temperatures, while simultaneously reducing the absolute concentration of individual buffers, thereby reducing or alleviating toxicity concerns. This report verifies that DIPSO, MOPS and HEPES, and their combinations, support embryo development. Therefore, utilization of bi- and tri-buffered media, containing a mixture of HEPES, MOPS or DIPSO, offers advantages compared with media containing HEPES or MOPS alone, and may be used for procedures such as oocyte retrieval, intracytoplasmic sperm injection, embryo transfer and cryopreservation.