MOUSE EMBRYO ASSAY (MEA): IT’S NOT WHAT YOU DO, IT’S THE WAY THAT YOU DO IT!

For quality control, manufacturers perform a Mouse Embryo Assay (MEA) on most medical devices used within IVF. The MEA, however, is not a standardized embryotoxicity assay and the normal endpoint is the percentage of embryos that have reached the expanded blastocyst stage (i.e., ≥80%) after 96 hours of culture. Recently, the FDA released a guideline to help shape the assay (https://www.fda.gov/media/127874/download). The purpose of this study was to demonstrate that how a MEA is performed, and the endpoints used, dictate the sensitivity of the assay. To investigate if determining the blastocyst cell number is a more sensitive endpoint than morphology, 1-cell mouse embryos (F1) were incubated in a medium containing 0.0, 0.002, 0.004 or 0.008% Triton X-100 for 96 hours. Groups of up to 10 embryos were cultured in 20μL under an oil overlay. The embryos that had reached the expanded blastocyst stage were recorded and then the number of cells in each blastocyst were determined using a nuclear stain. For testing various oils, 1-cell embryos were cultured in groups as above or individually in 2μL drops (to increase the surface area to volume). Blastocyst cell numbers were compared to the control using an unpaired t-test. At 0.002% Triton X-100, embryo development reached 86% expanded blastocyst. Blastocyst cell number was not significantly different to the control. A 0.004% solution supported 87% expanded blastocyst development. Blastocyst cell number was significantly decreased (p<0.0001). Expanded blastocyst development was 1% with a 0.008% solution. The table below shows that differences in oil quality can also be demonstrated by determining the blastocyst cell number. With Oil B the blastocyst cell number was significantly decreased following group culture (*p<0.02). The microdrop assay further increased the sensitivity of the MEA as Oil B only supported 15% expanded blastocyst development.Tabled 1Embryos (n)Exp. Blastocysts at 96 hours (%)Mean blastocyst cell number (sem)Control Oil (group)30100131 ±4Test Oil A (group)6098133 ± 3Test Oil B (group)6092117 ±4*Control Oil (microdrop)6088105 ±3Test Oil A (microdrop)6092101 ±3Test Oil B (microdrop)601584 ±10 Open table in a new tab MEA sensitivity can be modified by how it is performed. Using Triton X-100 demonstrates that cell number can detect a suboptimal culture component that would not be revealed by the ≥80% expanded blastocyst criteria. Furthermore, individual culture in a reduced volume appears to be a more suitable test for an oil overlay.