P-259 Intracytoplasmic sperm injection conducted within a micro 3D printed device that requires no holding pipette or vacuum reduces porcine oocyte invagination during microinjection

Abstract

Abstract Study question Can the use of a custom-designed and 3D printed microICSI™ device reduce the degree of oocyte invagination during intracytoplasmic sperm injection (ICSI)? Summary answer Invagination was significantly less in porcine oocytes when ICSI was performed in microICSI™ device compared to conventional ICSI. What is known already ICSI is a difficult procedure for embryologists to master, yet until recently, technically little had changed. In particular, the holding pipette (HP) with vacuum to hold the oocyte has not been surpassed. However, a source of shear-stress during ICSI of oocytes is the invagination of the zona pellucida and cytoplasm that occurs during microinjection, due to the pressure of the injection pipette against the small surface area of the HP. A recent study demonstrated microinjection within a two-piece 3D printed device. Here we evaluated a design refinement that displaced injection pressure more evenly across the oocyte to improve embryo quality. Study design, size, duration A control (conventional ICSI) and study group (microICSI™) were compared over three replicates investigating the level of invagination (3-point score, 1 is lowest and 3 is greatest) during ICSI. Incidence of cytoplasmic lysis was also recorded 24 h following ICSI. Participants/materials, setting, methods Porcine oocytes collected by follicle aspiration of abattoir-sourced ovaries were matured for 39-44 h in NCSU-17 medium supplemented with eCG, hCG, porcine follicular fluid, EGF and insulin. Mature oocytes, stripped of cumulus, were randomly allocated to either conventional ICSI or microICSI™. ICSI was performed, with treatment order randomised, and the injected oocytes were cultured in NCSU-17 culture medium at 39 C. Main results and the role of chance The microICSI™ device was modelled as a one-piece structure with contoured surfaces within a microwell array to support the oocyte during injection without the requirement for a holding pipette. Two-photon polymerisation 3D micro printing was used to print and assess multiple prototype geometries to optimise the support of the oocyte during ICSI. Over the three replicates (n = 10-20/treatment/replicate), the degree of invagination was consistently lower in the microICSI™ group compared with conventional ICSI (Mean ± SEM): Replicate 1: 1.40 ± 0.16 vs. 2.00 ± 0.19; Replicate 2: 1.20 ± 0.11 vs. 2.00 ± 0.13; Replicate 3: 1.42 ± 0.12 vs. 1.78 ± 0.15, with a paired t-test showing a significant difference between the two groups (p = 0.035). The incidence of lysis was not significantly different between the two treatments (ranging from 15-20% across both groups and varied between replicates). Limitations, reasons for caution The use of a porcine ICSI model may not sufficiently replicate what occurs with human ICSI as human oocytes experience greater levels of invagination (typically scored out of 4 rather than 3). Wider implications of the findings Reducing the invagination due to the injection process may reflect a less stressful procedure, with improvement to fertilization rates and embryo quality. Trial registration number not applicable