生物
标记法
细胞凋亡
末端脱氧核苷酸转移酶
吞噬作用
原位缺口末端标记
细胞生物学
凋亡DNA断裂
吖啶橙
DNA断裂
膜联蛋白
半胱氨酸蛋白酶
磷脂酰丝氨酸
分子生物学
程序性细胞死亡
生物化学
磷脂
膜
作者
Kristin White,Simonetta Lisi,Phani Kurada,Nathalie C. Franc,Peter Bangs
标识
DOI:10.1016/s0091-679x(01)66015-1
摘要
This chapter discusses the several techniques have been developed to label apoptotic cells in Drosophila tissues. Both acridine orange (AO) and the transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) technique can be used to assess apoptosis in the developing embryo and in the larval imaginal discs and nervous system. The molecular basis of AO staining is controversial but seems to involve specific retention of the dye in apoptotic cells. The TUNEL technique takes advantage of the DNA cleavage found in apoptotic cells. The free DNA ends targeted by terminal deoxytransferase (TdT), incorporates digoxygenin-labeled nucleotides. This chapter provides protocols for AO and TUNEL labeling, as well as for ultrastructural studies. The chapter discusses the other assays used for labeling apoptotic cells in Drosophila. Activation of caspases is a stereotypic feature of apoptotic cells and can be detected in situ with affinity-labeling techniques. The binding of Annexin V to exposed phosphatidylserine is another common technique used to label apoptotic cells. This chapter provides a protocol to assess embryonic phagocytosis of apoptotic cells. Phagocytosis also occurs later in development, a time when hemocytes are abundant and more accessible by bleeding.
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