Safe and efficient detection of egg maturity without cumulus cell removal by non-invasive tomography

Currently, eggs must be denuded to assess maturity. However, maintenance of the cumulus cell investment is critical to support oocyte quality during maturation. A novel tomography device using near infra-red light has been developed to detect the first polar body without removing cumulus cells. Our objective was to determine the effect of tomographic imaging for polar body detection on mouse oocyte developmental competence and subsequent embryo quality. Prospective research study. The experimental design consisted of three treatment groups; control eggs (C, standard IVF/IVC protocol; n=135), imaged eggs (I; n=45), and eggs that were treated identically to I but not exposed to tomography (NI; n=63). Two replicates were performed. In vivo matured mouse (outbred CF1, n=8 females) eggs were collected following ovarian stimulation. A subset of oocytes was randomly selected and immediately placed into fertilization medium containing sperm (C). The remaining eggs were placed into two imaging dishes, consisting of 50 μL drops of MOPS buffered medium under oil. Both dishes were moved to a heated stage on the imaging system. Eggs in one dish were imaged (I; 8-10 eggs/microdrops); oocytes in the second dish (NI) served as an environmental control. Presence or absence of the polar body was recorded for each egg. After imaging, both I and NI groups were placed into fertilization medium containing sperm. After IVF, 2PN zygotes were placed into sequential culture medium. It required 93 seconds on average to image each oocyte and determine if it contained a polar body (range, 40-150 sec/oocyte). Tomography evaluation revealed that 78% of the eggs were mature. After IVF, C had fewer (p<0.01) 2PN zygotes than either I or NI (60%, 87%, 76%, respectively). The percentage of mature eggs (78%) was slightly underestimated compared to the percentage of successfully fertilized zygotes (87%). There were no differences in blastocyst development or hatching between treatments (C, 64% and 57%; I, 67% and 69%; NI, 75% and 67%, respectively). There were no differences in ICM or total cell number between treatments, although NI tended (p=0.08) to have fewer TE cells than C (NI, 83.6±5.3; C, 100.8±6.1). The percentage of ICM cells was increased (p<0.05) in I (12.3±1.1%) compared to C (8.9±0.7%). The expression of 8/9 genes related to blastocyst viability (BMP15, DNMT3A, FOXO3A, GLUT1, GRP78, NANOG, PASG, PLAC8) did not differ between treatments, although expression of ATF4 was decreased (p<0.05) in I and NI compared to C. Assessment of cumulus enclosed mouse eggs to determine maturity using near infra-red tomography does not have any negative effects on fertilization, blastocyst development or embryo quality. This data suggests that tomography could be used to safely make clinical decisions about the most appropriate fate of each retrieved egg prior to cumulus removal, thereby improving quality of the oocyte cohort.