## 2 IVF Culture, Embryo Development, and Selection
### Stages of Embryo Development in IVF  
In the IVF process, the quality of fertilized eggs is highly important as the selection of a good fertilized egg could ensure a successful pregnancy rate [9; 16] before the embryo is transferred into the uterus. The fertilized egg develops into a human embryo (zygote) and can be presented in four stages, namely, a pronucleate oocyte (pronuclear), cleavage stage, morula, and blastocyst, as presented in Table 1, with respect to the hours post insemination and its expected features of each stage of development.  
Beside the developed embryo of day 0 to day 3, the morula has been least characterized for embryo grading due to the difficulty in assessing the evide  
\begin{table}
\begin{tabular}{l c c c} \hline \hline \multirow{2}{*}{**Stage**} & \multirow{2}{*}{**Timing (h) [17]**} & **Expected Features** & **Ideal Morphology** \\  & & **Developed [17]** & **Features/Visibility [18]** \\ \hline \multirow{3}{*}{Pronuclear (Day 0)} & \multirow{3}{*}{\(17\pm 1\)} & \multirow{3}{*}{Pronucleate oocyte} & (i) NPB 1 \textless{} 3 \\  & & & (ii) NPB always polarized \\  & \(23\pm 1\) to \(26\pm 1\) & & Up to 20\% may be at & (i) Mononucleated blastomeres \\ Cleavage (Day 1) & (post ICSF 2), \(28\pm 1\) & & or reach the & (ii) Equal cell sizes \\  & (post IVF) & & two-cell stage & (ii) \textless{}20\%/s fragmentation \\  & & & (i) Mononucleated blastomeres \\ Cleavage (Day 2) & \multirow{3}{*}{\(44\pm 1\)} & \multirow{3}{*}{Four-cell stage} & (ii) Equal cell sizes \\  & & & (iii) \textless{}20\%/s fragmentation \\  & & & (i) Mononucleated blastomeres \\ Cleavage (Day 3) & \multirow{3}{*}{\(68\pm 1\)} & \multirow{3}{*}{Eight-cell stage} & (ii) Equal cell sizes \\  & & & (iii) \textless{}20\%/s fragmentation \\  & & & (iv) at least seven blastomeres \\ Morula (Day 4) & \multirow{3}{*}{\(92\pm 2\)} & \multirow{3}{*}{Compaction volume} & (i) Compact cells (by increase in \\  & & & embryo and ZP space) \\  & & & (ii) Lack of fragments \\  & & & (i) Expanded blastococcal cavity \\ Blastocyst & \multirow{3}{*}{\(116\pm 2\)} & \multirow{3}{*}{Fully expanded, through-to-hatched} & (ii) composed of many inner \\  & & & cells mass \\ (Day 5/6/7) & & & (iii) Cohesive epithelium cells at TE \\  & & & (tropbedoderm) \\  & & & (iv) Zona Pellucida (ZP) thinning \\ \hline \hline \end{tabular}  
* NPB = nucleolar precursor bodies, 2 ICSI = intracytoplasmic sperm injection.  
\end{table}
Table 1: Embryo stages and morphological features in IVF process from pronuclear to blastocyst stage of development at each time point.  
fertilized embryo and the less well-defined blastomere boundaries [17; 19]. Due to the ideal morphological features, blastocysts at day 5 are reported to have higher implantation potential compared to other embryo stages [17]. Hence, morphology-based embryo selection to quantify a good and high-quality blastocyst prior to embryo grading has become more critical to improve the IVF success rate. Meanwhile, delayed blastocyst development at day 7 or 8 is considered a poor prognosis for implantation [20] and they are routinely discarded, since the proportion of top-quality blastocysts are lower at day 7 [7]. Consequently, there are many detailed assessment and grading schemes have been devised for grading the quality of embryos from the pronuclear stage [21], cleavage stage [22], morula stage [19], and blastocyst stage [23; 24].