Abstract Study question 3-dimensional quantitative assessment of the inner cell mass (ICM) is more effective at selecting viable embryos than the widely used Gardner grading system? Summary answer 3-dimensional quantitative assessment of the ICM is significantly more predictive of implantation and live birth than the widely used Gardner grading of the ICM What is known already ICM assessment forms a key part of most widely used blastocyst grading systems. Grading under these systems, however, is often qualitative, subjective, and subject to inter/intra-operator variability. To reduce subjectivity in ICM grading, it has been suggested that quantitative criteria are used. Previous studies have focused on the assessment of such parameters (e.g. ICM area) in 2 dimensions. However, such approaches fail to account for the various embryo orientations which could lead to different measurements. Therefore, quantitative 3-dimensional analysis of the ICM is needed to improve the reliability and consistency of grading. Study design, size, duration This was a retrospective analysis of 153 randomly selected tEB embryos from two clinics. All data was captured on Embryoscope between 2018 and 2020. Data for maternal age, blastocyst grade, blastocyst ploidy, implantation and live birth rates were also recorded. Participants/materials, setting, methods Embryos were imaged at 7+ focal planes and in-focus sections of each ICM were annotated using ImageJ. These sections were combined into a 3D model using Blender, and sphericity (ψ) computed. Embryos were categorised as spheroid (ψ ≥ 0.8), ellipsoid (ψ < 0.8), crescent and irregular. Chi-squared tests were performed, comparing classification and clinical outcome. The predictive power of the system was compared to the Gardner scale (spheroid/ellipsoid classifications and A grades considered predictive of a positive outcome respectively). Main results and the role of chance Most ICM were either ellipsoid (48%) and spheroid (37%) rather than crescent (6%) or irregular (8%, p < 0.001). ICM 3D classification was associated with implantation, X2(3, N = 119)= 11.10, p = 0.01 and live birth, X2(3, N = 119)= 10.78, p = 0.01. Blastocysts with ellipsoid (70%, 28/40) and spheroid (58%, 18/31) shaped ICM had the greatest chances of implantation, whilst those with crescent (40%, 6/15) and irregularly shaped ICM (33%, 11/33) had lower chances of implantation (p < 0.011). Similarly, blastocysts with ellipsoid (58%, 23/40) and spheroid (48%, 15/31) shaped ICM had the greatest chances of live birth, whilst those with crescent (33%, 5/15) and irregularly shaped ICM (21%, 7/33) had lower chances of live birth (p < 0.013). 3D assessment of ICM outperformed Gardner ICM grading with regards to prediction of implantation (accuracy 64.7% vs 49.6%, area-under-the-curve 0.642 vs 0.486, precision 64.8% vs 51.9%, sensitivity 73% vs 65.1%, specificity 55.4% vs 32.1%, F1 score 68.7% vs 57.8%, 3D vs Gardner) and live birth (accuracy 62.2% vs 50.4%, area-under-the-curve 0.641 vs 0.531, precision 53.5% vs 44.3%, sensitivity 76% vs 70%, specificity 52% vs 36.2%, F1 score 62.2% vs 54.3%, 3D vs Gardner). Limitations, reasons for caution Manually rendering the 3D images for each ICM was time consuming, making it impractical for clinical application. Further work is now focusing on automating this process using image recognition and augmented reality. Wider implications of the findings This is a novel ICM assessment that provides an improved perspective of the 3-dimensionality of the ICM which is more consistent and effective than traditional methods of embryo assessment, potentially contributing towards improving embryo selection efficacy and time to pregnancy. Trial registration number none