Oocyte activating physical method and fertilization potential evaluation index thereof
An oocyte and evaluation index technology, applied in the field of embryo engineering, can solve the problems of cell damage, excessive energy, poor repeatability, etc.
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Embodiment 1
[0025] Example 1: Activation of rabbit oocytes
[0026] 1. Experimental animals
[0027] Sexually mature female Japanese big-eared rabbits aged 6-15 months and weighing 2.5-3.5 kg were purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.
[0028] 2. Experimental reagents
[0029] Pregnant Horse Serum (PMSG Sigma)
[0030] Human chorionic gonadotropin (hCG Sigma)
[0031] Plasma Substitute (SSS Irvine Scientific)
[0032] Hyaluronidase (Sigma)
[0033] Modified Human Fallopian Tube Fluid (mHTF Irvine Scientific)
[0034] Human tubal fluid (HTF Irvine Scientific)
[0035] 3. Oocyte Preparation and Culture
[0036] First, the neck skin of female rabbits was disinfected with iodine cotton balls, and PMSG (150IU / rabbit) was subcutaneously injected into the neck. After 48 hours, the ear edge was sterilized with alcohol cotton balls, and hCG (75IU / a) was injected into the ear vein for superovulation. 24h after the injection of hCG, the female rabbits w...
Embodiment 2
[0044] Example 2: Induction of calcium ion oscillations in mouse oocytes
[0045] 1. Experimental animals
[0046] Sexually mature female Kunming mice aged 6-8 weeks were purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.
[0047] 2. Experimental reagents
[0048] Pregnant Horse Serum (PMSG Sigma)
[0049]Human chorionic gonadotropin (hCG Sigma)
[0050] Plasma Substitute (SSS Irvine Scientific)
[0051] Hyaluronidase (Sigma)
[0052] Modified Human Fallopian Tube Fluid (mHTF Irvine Scientific)
[0053] Human tubal fluid (HTF Irvine Scientific)
[0054] Fluo-4AM / Calcium Fluorescent Probe (Dojindo Laboratories)
[0055] 3. Oocyte Preparation and Culture
[0056] PMSG (10IU / mice) was injected intraperitoneally to female mice, and hCG (10IU / mice) was injected intraperitoneally 48 hours later for superovulation. After 15-19 hours of hCG injection, the mice were sacrificed by cervical dislocation, and the fallopian tubes were exposed and removed un...
Embodiment 3
[0061] Example 3: Evaluation of the fertilization potential of A23187-activated mouse oocytes
[0062] 1. Experimental animals
[0063] Sexually mature female Kunming mice aged 6-8 weeks were purchased from Beijing Weitong Lihua Laboratory Animal Technology Co., Ltd.
[0064] 2. Experimental reagents
[0065] Pregnant Horse Serum (PMSG Sigma)
[0066] Human chorionic gonadotropin (hCG Sigma)
[0067] Plasma Substitute (SSS Irvine Scientific)
[0068] Hyaluronidase (Sigma)
[0069] Modified Human Fallopian Tube Fluid (mHTF Irvine Scientific)
[0070] Human tubal fluid (HTF Irvine Scientific)
[0071] Fluo-4AM / Calcium Fluorescent Probe (Dojindo Laboratories)
[0072] Calcium ionophore A23187 (A23187Sigma)
[0073] 3. Oocyte Preparation
[0074] PMSG (10IU / mice) was injected intraperitoneally to female mice, and hCG (10IU / mice) was injected intraperitoneally 48 hours later for superovulation. After 15-19 hours of hCG injection, the mice were sacrificed by cervical dislo...
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