since 1978 , when the first baby conceived in vitro was born , in vitro fertilization ( ivf ) has become a major method for treating infertility . although ivf procedures have been greatly improved over the years , their efficiency , as defined by live birth rate , is still only about 30 - 40% .
consequently , many couples have to undergo treatment several times before they succeed and this increases emotional and financial costs , as well as additional health risks for women
. the efficiency of ivf can be increased by transferring multiple embryos in a single cycle , but this may lead to multiple - pregnancies , and consequently serious health complications for mothers and babies .
there is , therefore , a strong need for single embryo transfers , and so the development of reliable methods of selecting embryos is crucial . here
, we provide a short review of current techniques for embryo assessment , with a focus on new ways of distinguishing high - quality embryos based on recent advances in time - lapse imaging technology .
visual assessment of embryo morphology is the most traditional and popular method of embryo selection .
several parameters can be assessed at different developmental stages , providing valuable information about the quality of embryos .
embryos can be graded according to the morphology of their pronuclei on day 1 after fertilization [ 4 - 6 ] , the number and shape of blastomeres and degree of fragmentation on day 2 or 3 [ 7 - 9 ] , or by the morphology of the blastocyst at day 5 or 6 [ 10 - 12 ] . depending on the procedure , embryos are evaluated at one or several developmental stages . although morphological assessment is inexpensive and easy to implement in a clinical environment , it has its drawbacks .
moreover , even when such expertise is available , the technique is not always accurate : low - graded embryos often prove to have high developmental potential and can develop to term .
therefore , there has been a drive to develop an alternative method of embryo evaluation that provides more detailed information about the developmental status of the embryo and , most importantly , is quantitative and objective .
it uses polymerase chain reaction ( pcr)-based techniques to diagnose specific genetic mutations   or fluorescence in situ hybridization ( fish ) to diagnose chromosomal abnormalities or to sex the embryos for patients carrying x - chromosome - linked diseases [ 14 - 16 ] .
preimplantation genetic diagnosis can be applied to embryos at different stages : zygotes ( polar body biopsy ) , cleavage stage embryos ( blastomere biopsy ) or blastocysts ( trophectoderm biopsy ) .
an alternative approach is preimplantation genetic screening , which was developed to improve ivf outcomes in mothers of advanced age , those presenting with repeated miscarriages or implantation failure , or in cases of severe male factor infertility .
the classic form of preimplantation genetic screening , involving fish assessment of a limited number of chromosomes in one blastomere of a cleavage stage embryo , has been shown to be ineffective and has been gradually replaced by technologies based on comparative genomic hybridization ( cgh ) or single nucleotide polymorphism ( snp ) arrays . both of these approaches allow the whole embryonic genome to be analysed and , as for preimplantation genetic diagnosis , used at different embryonic stages .
changes in pyruvate or glucose concentration in the culture medium can reflect the energy metabolism of the embryo , although their usefulness as a tool to predict the embryo 's quality is not clear [ 19 - 23 ] . on the other hand , oxygen consumption [ 24 - 27 ] and amino - acid turnover [ 28 - 30 ]
recently , the analysis of single metabolites has been gradually replaced by a broader approach : metabolomic or protein secrotome profiling , which can provide a complete picture of an embryo 's metabolism and gene expression patterns .
although many reports have shown a correlation between metabolic status of the embryo and its viability , establishing its potential value for the ivf clinics , for the moment these techniques remain difficult to implement in a clinical environment .
this is because metabolomic or secreted protein analyses involve spectroscopic / spectrometric and chromatographic techniques , which currently require expensive equipment and highly skilled staff .
the method described by wong et al . involves time - lapse imaging of embryos every 5 min for several hours ( from 1- to 4-cell stage ) . in comparison , the method established by ajduk et al . requires a much shorter period of recording ( 2.5 hours for mouse embryos ) but with images taken every 10 seconds .
due to recent advances in non - invasive time - lapse imaging established in mouse embryos , we can now follow the dynamics of embryo divisions and other fertilization - triggered events and correlate them with the developmental potential of the embryos .
the last two years have seen two studies employing this technique in a very different way .
a team at stanford university has shown that the timing and synchrony of the first two embryonic divisions are predictive of developmental potential of human embryos ( figure 1 ) .
the authors reported that embryos with a very long first cytokinesis , with a prolonged or an atypically short interval between first and second division or with highly asynchronous divisions of two - cell blastomeres , fail to reach the blastocyst stage .
this accords with previous observations , in which timely pronuclear formation and subsequent first cleavage correlated with higher quality of human embryos [ 34 - 36 ] .
a completely different approach has been developed by our team at university of cambridge working collaboratively with teams at oxford and cardiff universities ( figure 2 ) . in this work
, we showed that fertilization of mouse eggs triggers abrupt , repetitive cytoplasmic movements that correlated with ca oscillations ( also triggered by sperm ) and depended on the functionality of the cytoskeleton .
embryos that showed very frequent increases in cytoplasmic movements ( indicating very frequent ca oscillations ) and low cytoplasmic speeds in the intervening periods ( reflecting low quality of the actin cytoskeleton ) were three - fold less successful in developing to pups than embryos displaying average values of these parameters .
although both methods still have to be tested in a clinical environment and subjected to randomised controlled trials to demonstrate improved live birth outcomes , they offer great hope for more reliable assessment of embryonic quality .
mouse eggs are subjected to time - lapse imaging ( 1 frame every 10 seconds for 2.5 hours ) immediately after fertilization .
acquired images are analysed by the particle image velocimetry method that follows patterns of contrasts between subsequent images and calculates how they move .
the sum of all displacement vectors calculated for the zygote in a given time - point ( i.e. mean cytoplasmic speed ) is plotted over time .
mean interval between the fast movements ( in red ) and mean speed in periods inbetween the fast movements ( mean basal speed , in blue ) are indicative of developmental potential of the embryo .
research carried out over the past three decades has provided a broad repertoire of possible improved embryo selection methods . some of them , such as evaluation of pronuclear morphology , preimplantation genetic diagnosis and preimplantation genetic screening ,
others ( e.g. analysis of metabolites , secreted proteins or cytoplasmic flows ) concentrate on the cytoplasmic component , analysing the quality of embryo metabolism , the cytoskeleton or ca homeostasis .
this can be achieved for instance by following the timing of embryonic cell divisions , as their duration and synchrony can be affected by improper chromosome segregation , cytoskeletal properties and energy levels .
an alternative approach could combine preimplantation genetic screening with the analysis of embryo metabolism or cytoplasmic movements .
combination of genetic testing of the polar body and examination of the cytoplasmic flows is especially promising , as it would provide information about embryo quality within several hours after fertilization , and , therefore , significantly quicken the selection process ( figure 1 ) .
indeed , recent publications suggest that a shorter period between in vitro fertilization and embryo transfer may be beneficial , as prolonged in vitro culture of embryos alters their epigenetic modifications and gene expression . through a combination of these new methods ,
it is reasonable to expect that embryo selection for ivf will become much more reliable in the coming years .
the method of assessing embryo developmental potential based on analysis of the cytoplasmic movements is the subject of a patent application filed in the u.s . by magdalena zernicka - goetz and anna ajduk ( us
 despite many recent advances in the field of reproductive biology and medicine , the efficiency of in vitro fertilization procedures remains relatively low . 
 there is a need for a reliable and non - invasive method of embryo selection to ensure that only embryos with the highest developmental potential are chosen for transfer to mothers - to - be . here , 
 we compare various methods currently used for assessing embryonic viability , such as examination of embryonic morphology , quality of the genetic material , or metabolism . 
 additionally , we discuss novel procedures for embryonic assessment based on advanced time - lapse imaging techniques , which show great promise and may lead to increased in vitro fertilization efficiencies . 
Introduction
Static assessment of embryo morphology by light microscopy
Preimplantation genetic diagnosis and screening
Metabolomics and proteomics in screening
Dynamic assessment of embryonic development by time-lapse imaging
Conclusion
Competing interests