摘要
The brilliant cresyl blue (BCB) test determines the activity of glucose-6-phosphate dehydrogenase (G6PDH); the activity of this enzyme is greatest in growing oocytes, but it declines as oocytes mature. The objective was to develop and evaluate this test for assessing development of buffalo oocytes (to select developmentally competent oocytes for increased in vitro embryo production). Oocytes were exposed to BCB stain diluted in mDPBS (DPBS with 0.4% BSA) for 90 min at 38.5 °C in a humidified air atmosphere; those with or without blue coloration of the cytoplasm were designated as BCB+ and BCB−, respectively. In Experiment 1, oocytes were exposed to 13, 26, or 39 μM BCB. There were fewer BCB+ oocytes after exposure to 13 μM BCB (10%) than after exposure to 26 or 39 μM BCB (57.2 and 61.8%; P < 0.05), but there was no significant difference among treatments for blastocyst production rate. In Experiment 2, the diameter of BCB+ oocytes (144.4 ± 4.2 μm; mean ± S.E.M.) was higher (P < 0.05) than that of BCB− oocytes (136.8 ± 4.6 μm). In Experiment 3, oocytes were allocated into three groups: control (immediately cultured); holding-control (kept in mDPBS for 90 min before cultured); and treatment-incubation (incubated with 26 μM BCB). After IVM, oocytes were fertilized in vitro and cultured on an oviductal monolayer. The nuclear maturation rate was higher (P < 0.05) in BCB+ (86.2%), control (83.4%) and holding-control (82.6%) oocytes than BCB− (59.2%) oocytes. The BCB+ oocytes yielded more blastocysts than control or holding-control oocytes (33.4, 20.2, and 21.0%, P < 0.05); blastocyst development was lowest in BCB− oocytes (5.2%). In conclusion, staining of buffalo oocytes with BCB before IVM may be used to select developmentally competent oocytes for increased in vitro embryo production.
摘要译文
灿烂甲酚蓝（BCB）试验测定葡萄糖-6-磷酸脱氢酶（G6PDH）的活性;这种酶在卵母细胞生长中的活性最大，但随着卵母细胞的成熟，其活性下降。目的是开发和评估这个测试来评估水牛卵母细胞的发育（选择发育上有能力的卵母细胞来增加体外胚胎生产）。将卵母细胞暴露于在mDPBS（含有0.4％BSA的DPBS）中稀释的BCB染色液中，在38.5℃，潮湿空气气氛中孵育90分钟;具有或不具有蓝色着色的细胞质分别命名为BCB +和BCB-。在实验1中，将卵母细胞暴露于13,26或39μMBCB。接触13μMBCB（10％）后的BCB +卵母细胞比暴露于26或39μMBCB（57.2和61.8％; P 0.05）但囊胚产生率处理间差异不显着。在实验2中，BCB +卵母细胞的直径（144.4±4.2μm;平均值±S.E.M.）较高（P 0。05）比BCB-卵母细胞（136.8±4.6μm）。在实验3中，将卵母细胞分成三组：对照（立即培养）;保持对照（在培养前保持在mDPBS中90分钟）;和处理孵育（用26μMBCB孵育）。 IVM后，将卵母细胞体外受精并在输卵管单层上培养。 BCB +的核成熟率较高（P 0.05）。2％），对照组（83.4％）和保持对照组（82.6％）卵母细胞比BCB-（59.2％）卵母细胞。 BCB +卵母细胞比对照组和控股卵母细胞产生更多的囊胚（33.4％，20.2％和21.0％，P 0.05）;在BCB-卵母细胞中胚泡发育最低（5.2％）。结论是，在IVM之前用BCB染色水牛卵母细胞可用于选择发育上有活力的卵母细胞以增加体外胚胎生产。
B.M.Manjunatha[b];P.S.P.Gupta[a];M.Devaraj[b];J.P.Ravindra[a];S.Nandi[a];. Selection of developmentally competent buffalo oocytes by brilliant cresyl blue staining before IVM[J]. Theriogenology, 2007,68(9): 1299-1304