Carroll David J, Hua Wei
Department of Biological Sciences, Florida Institute of Technology, Melbourne, FL, USA.
Methods Mol Biol. 2009;518:57-66. doi: 10.1007/978-1-59745-202-1_5.
The starfish oocyte has proven useful for studies involving microinjection because it is relatively large (190 mum) and optically clear. These oocytes are easily obtained from the ovary arrested at prophase of meiosis I, making them useful as a model system for the study of cell cycle-related events. In this chapter, a method for combining microinjection with immunoblotting of single cells is described. Individual starfish oocytes are injected, removed from the microinjection chamber, and analyzed by immunoblotting for the dual-phosphorylated form of mitogen-activated protein kinase (MAPK). This method will allow for experiments testing the regulation of MAPK in single cells and for the manipulation of these cells by a quantitative microinjection technique.
海星卵母细胞已被证明适用于涉及显微注射的研究，因为它相对较大（190微米）且光学透明。这些卵母细胞很容易从停滞在减数分裂I前期的卵巢中获得，使其成为研究细胞周期相关事件的有用模型系统。在本章中，描述了一种将显微注射与单细胞免疫印迹相结合的方法。对单个海星卵母细胞进行注射，从显微注射室中取出，然后通过免疫印迹分析有丝分裂原激活蛋白激酶（MAPK）的双磷酸化形式。该方法将允许进行测试单细胞中MAPK调节的实验，并通过定量显微注射技术对这些细胞进行操作。
