Binding and fusion of Hoechst-labelled sperm to ZP-free oocytes matured in the presence of 43 µM or 100 µM LA (fresh and vitrified)
; Suqueli García, María Florencia ; Manrique, Rodrigo Javier ; Buschiazzo, Jorgelina Otros responsables
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Publicador:
Consejo Nacional de Investigaciones Científicas y Técnicas
Fecha de depósito:
3/06/2026
Fecha de creación:
2025-2025
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The file contains the folowing columns : FRESH: non-vitrified oocytes. VITRIFIED: oocytes vitrified using a surface device (Cryotech®). CONTROL : oocytes matured without LA. 43 µM LA: oocytes matured in 43 µM LA. 100 µM LA: oocytes matured in 100 µM LA. # experiment: number of experiments performed (within each treatment the rows correspond to oocytes evaluated).
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DATA FROM: Binding and fusion of Hoechst-labelled sperm to ZP-free oocytes matured in the presence of 43 µM or 100 µM LA (fresh and vitrified). LA: linoleic acid; ZP: zona pellucida In vitro fertilization of ZP-free oocytes: Methodology Cumulus cells were removed by a brief exposure to hyaluronidase (1 mg/mL). Only oocytes showing a visible polar body after in vitro maturation were used in this assay. The ZP was digested with 1 mg/mL pronase (protease from Streptomyces griseus) under visual monitoring. The ZP-free oocytes were rapidly washed in HSOF-PVA and kept at 38.5°C under 5% CO2 in humidified air for 2 recovery h. Next, ZP-free oocytes were inseminated in 100 μL drops of IVF-SOF medium supplemented with 12 μg/mL heparin. Co-incubation with sperm at a final concentration of 1 × 10^5 spermatozoa/mL was performed for 16 h at 38.5°C under 5% CO2 in humidified air. After insemination, the oocytes were washed through 3 drops by the same person using the same thin-bore pipette. Oocytes were then stained with 5 μM bisBenzimide Hoechst 33342 for 5 min at room temperature and mounted for analysis under an epifluorescence microscope (Nikon TE-300; Nikon, Tokyo, Japan). Digital photographs were taken with a DSfi1 camera (Nikon, Tokyo, Japan) connected to the microscope.
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INTA_CRBsAsSur_EEABalcarce_Rios_GL_Binding_fusion_sperm_ZP-free_oocytes.xlsx
http://hdl.handle.net/20.500.12123/22579 (repositorio del INTA)
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