Immunochemical visualization and quantitation of cyclic AMP in single cells

被引:19
作者
Wiemelt, AP
Engleka, MJ
Skorupa, AF
McMorris, FA
机构
[1] WISTAR INST ANAT & BIOL, PHILADELPHIA, PA 19104 USA
[2] UNIV PENN, DAVID MAHONEY INST NEUROL SCI, PHILADELPHIA, PA 19104 USA
关键词
D O I
10.1074/jbc.272.50.31489
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Adenosine 3':5'-cyclic monophosphate (cAMP) is a key second messenger in signaling pathways governing many cellular processes. To define the subcellular localization and relative abundance of cAMP, we developed a novel immunochemical approach based on acrolein fixation to visualize cAMP within cells. We describe here the fixation and immobilization of cAMP within cells and the production of specific, high titer polyclonal antibodies that recognize cAMP. Relative levels of cAMP immunofluorescence were quantitated in glial cells (oligodendrocytes, astrocytes, Schwann cells, and glioma cells) that were either untreated or treated with activators of endogenous adenylyl cyclase to raise cAMP levels, In treated cells, cAMP immunofluorescence is strongly localized in the perinuclear cytoplasm.
引用
收藏
页码:31489 / 31495
页数:7
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