Supplementatry Table 1 Outline of the national grading scheme for cleavage and blastocyst stage embryos (adapted from Cutting et al., 2008).

Parameter / Grade
Cleavage-stage embryo grading
Blastomere number / The number to denote how many blastomeres are present at the time of observation
Blastomere size / 4 = regular, even division
3 = <20% difference in blastomere diameter
2 = 20-50% difference
1 = >50% difference
Fragmentation / 4 = <10% fragments by volume
3 = 10-20%
2 = 20-50%
1 = >50%
Blastocyst stage embryo grading
Expansion status / 1= early blastocyst; blastocoel less than half the volume of the embryo, little or no expansion in overall size, zonapellucida (ZP) still thick
2 = Blastocyst; blastocoel cavity more than half the volume of the embryo, some expansion in overall size, ZP beginning to thin
3 = Full blastocyst; blastocoel completely fills the embryo
4 = Expanded blastocyst; blastocoel volume now larger than that of the early embryo. ZP very thin.
5 = Hatching blastocyst; trophectoderm has started herniated through the ZP
6 = Hatched blastocyst; the blastocyst has evacuated the ZP
Inner cell mass / A = Inner cell mass prominent, easily discernable and consisting of many cells, cells compacted and tightly adhered together
B = Cells less compacted so larger in size, cells loosely adhered together, some individual cells may be visible
C = Very few cells visible, either compacted or loose, may be difficult to completely distinguish from trophectoderm
E = No inner cells mass cells discernable in any focal plane
Trophectoderm / A = Many small identical cells forming a continuous trophectoderm layer
B = Fewer, larger cells, may not form a completely continuous layer
C = Sparse cells, may be very large, very flat or appear degenerate

Supplementary Table 2 Optimum time ranges and ESA grade assignment using the hierarchical embryo selection algorithm employed at the study site during the investigation.

Parameter / Optimum time range
Tier 1 / s2 / 0- 1.7h
Tier 2 / cc3 / 9.5-11.4h
Tier 2 / t5 / 43.46-51.04h
ESA grade / Tier 1 / Tier 2 / Tier 3
A+ / Fulfilled / Fulfilled / Fulfilled
A- / Fulfilled / Fulfilled / Not fulfilled
B+ / Fulfilled / Not fulfilled / Fulfilled
B- / Fulfilled / Not fulfilled / Not fulfilled
C+ / Not fulfilled / Fulfilled / Fulfilled
C- / Not fulfilled / Fulfilled / Not fulfilled
D+ / Not fulfilled / Not fulfilled / Fulfilled
D- / Not fulfilled / Not fulfilled / Not fulfilled
s2 = time between 3-cell and 4-cell. cc3 = time between 4-cell and 5-cell. t5 = time to 5-cell. Depending on whether the embryo fulfils the criteria set out in each tier (i.e. falls within the optimum time frames) it is assigned a grade from A+ to D-.

Supplementary Table 3Definitions of embryo quality used to classify embryos as good, average and poor quality based on ACE/BFS embryo grading guidelines (Cutting et al., 2008).

Good quality embryos (GQE) / Cleavage-stage embryos with even blastomeres (<20% difference in diameter) and <20% fragmentation
Blastocyst embryos with prominent and compact inner cell mass and many cells forming a cohesive epithelium
Average quality embryos (AQE) / Cleavage-stage embryos with 2050% difference in cell diameter and/or 2050% fragmentation
Blastocyst stage embryos with easily discernable inner cell mass with many cells that are loosely grouped together
Poor quality embryos (PQE) / Cleavage-stage embryo with >50% difference in blastomere diameter and/or >50% fragmentation
Blastocyst stage embryos with few cells forming the inner cell mass and very few cells making up the trophectoderm

Supplementary Table 4Embryo transfer baseline information for each abnormal embryo phenotype, including the total number of transfers, the number of single embryo transfers (SET), double embryo transfers (DET), cleavage-stage transfers and blastocyst-stage transfers.

Abnormal embryos
transferred (n) / Total transfers (n) / SET
(n) / DET (n) / Cleavage stage transfers (n) / Blastocyst stage transfers (n)
TDC / 1 / 1 / 1 / 0 / 0 / 1
FDC / 6 / 6 / 6 / 0 / 0 / 6
UDC / 16 / 15 / 14 / 1 / 5 / 10(1xDET)
Total DC / 23 / 22 / 21 / 1 / 5 / 17 (1xDET)
RC / 9 / 8 / 7 / 1 / 2 (1xDET) / 6
AC / 4 / 3 / 2 / 1 / 1 / 2 (1xDET)
CC / 48 / 37 / 26 / 11 / 20 (5xDET) / 17 (6xDET)
CL / 2 / 2 / 2 / 0 / 0 / 2
Overall / 86 / 72 / 58 / 14 / 28 / 44

Supplementary Table 5 Clinical pregnancy rates for all transfers including at least one abnormal embryo.

Transfers including an abnormal embryo (n) / Transfers including an abnormal embryo resulting in presence of fetal heart (n) / Clinical pregnancy rate (%)
DC / 44 / 4 / 9.1
RC / 13 / 0 / 0
AC / 6 / 1 / 16.7
CC / 74 / 2 / 2.7
CL / 5 / 0 / 0
Overall / 142 / 7 / 4.9