From a Backup Technology to a Strategy-Outlining Approach

The Success Story of Cryopreservation

Gábor Vajta; Anikó Reichart; Filippo Ubaldi; Laura Rienzi


Expert Rev of Obstet Gynecol. 2013;8(2):181-190. 

In This Article

Selection of the Best Technique

An informal survey has been performed to summarize the achievements over the past 6 years. A PubMed search performed on 14 April 2012 with a starting publication date of 1 January 2006 and closing date of 31 December 2011 for 'vitrification' or 'vitrified' resulted in 996 hits. Based on the title and abstract, publications dealing with human oocytes and embryos were individually selected. Reviews were excluded, except for those publishing data not fully included in the collected research papers. Eventually, 93 papers were collected and analyzed.

The first conclusion that can be drawn from these publications was that vitrification was found to be at least as efficient as traditional freezing. Better survival and developmental rates were experienced for oocytes in four and seven publications, respectively. Results were identical in one and two publications, respectively, while none of the 93 papers found a better outcome after cryopreservation with traditional slow-rate freezing. Vitrification was found to create less morphological and DNA damage than traditional freezing in human oocytes.[25] Two systematic reviews and meta-analyses, based on six and three papers, respectively, also found superior results achieved with vitrification.[26,27] Although the number of direct comparisons is relatively low, it is in perfect accordance with findings in other mammalian species. In some papers, in vitro and especially in vivo developmental rates were comparable between vitrification and traditional freezing; however, survival rates after both blastocyst and oocyte vitrification were significantly higher, approaching 94–100%, while this value was approximately 70 and 80% when traditional freezing was used.

Vitrification was also found to be harmless regarding the long-term consequences. Births were reported in 26 publications, including 11 after oocyte and 15 after embryo cryopreservation. None of these papers (including two systematic reviews[28,29]) found an increase in stillbirths or subsequent developmental of abnormalities. A recent study based on 6623 delivered singletons found an 85-g increase in the average birthweight, but no increase in birth defects and perinatal mortality rates after embryo vitrification compared with freshly transferred controls.[30] It should be noted that, according to a retrospective review, increased birthweight was found to be typical in singletons after cryopreservation regardless of the method used.[31] The same review also states that children born after embryo cryopreservation are healthy and have perinatal outcomes similar to their naturally conceived peers as well as better perinatal outcomes than children from fresh-embryo transfer.[31]

None of the 93 publications have reported in utero infections after oocyte or embryo cryopreservation – in full accordance with a wider survey that found no infections in human and domestic animal embryology attributable to any cryopreservation procedure.

Wherever applicable, some technical details were also collected and summarized. All except for one study used some forms of minimum-volume vitrification techniques; the vast majority of work was performed with open systems, that is, based on direct contact between the sample and liquid nitrogen (70 and six studies for open and closed systems, respectively). Closed systems used for blastocyst cryopreservation resulted in comparable results to open systems (four vs 19 papers, respectively); however, for metaphase II (MII) oocytes, only two papers presented data with closed-system vitrification, these data were seriously compromised compared with those presented in 23 publications referring to the use of open systems. Open systems resulted in less ultrastructural damage in MII human oocytes.[32] Ethylene glycol was the most frequently used permeable cryoprotectant, usually together with DMSO (55 papers), and propylene glycol replaced DMSO in four papers, all dealing with blastocyst cryopreservation and with lower survival and developmental tendencies.

Neither statistical analyses nor sound conclusions can be made from these diverse data, where variables surpassed constant parameters and conditions. However, the high amount of similar outcomes obtained in different laboratories by using the same technique may outline some tendency. Due to the lack of a sufficient number of prospective randomized studies comparing different methods, these tendencies may allow some orientation for outlining the best available technique for both oocyte and blastocyst vitrification; however, the best seems to be the minimum volume (≤1 µl), open system, with ethylene gycol and DMSO as the permeable cryoprotectants.