Paper Title
Assessment of the Impact of mice Superovulation and in Vitro Fertilization on Blastocyst Quality

Abstract
Transgenic mice are indispensable in biomedical research, relying heavily on assisted reproductive technologies (ARTs) for their generation and maintenance. Superovulation protocols play a crucial role in maximizing oocyte yield, enhancing the production of embryos for genetic modification. However, the impact of superovulation and in vitro culture on embryo quality remains a concern, potentially affecting early development and ART success rates. This study aimed to evaluate the quality of blastocysts produced from superovulation-induced mice, followed by in vitro fertilization (IVF) under defined culture conditions, with the intent of assessing the efficacy of the protocols routinely used in our laboratory. Specifically, we investigated whether these procedures affected crucial parameters linked to embryo quality, such as mitochondrial activity, phosphorylated-AKT (p-AKT) signaling, and S100 protein expression. The presence and activity of these markers were assessed using immunocytochemical staining techniques. The results indicated that mitochondrial activity, p-AKT signaling, and S100 protein expression were maintained at levels comparable to those observed under optimal physiological conditions. Mitochondrial activity was consistent, supporting adequate energy production required for cell division and differentiation. Likewise, the expression of p-AKT and S100 protein indicated that the regulatory pathways for cellular signaling and calcium homeostasis remained unaffected. These findings suggest that the applied IVF protocol, along with the specific culture conditions, did not compromise the intrinsic quality of the resulting blastocysts. Keywords - Blastocyst, Embryo Development, in Vitro Fertilization, Superovulation.