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对经微波固定的细胞进行免疫细胞化学分析显示，环磷酸腺苷（cAMP）或环磷酸鸟苷（cGMP）的亚细胞积累模式存在快速且激动剂特异性的变化。

Immunocytology on microwave-fixed cells reveals rapid and agonist-specific changes in subcellular accumulation patterns for cAMP or cGMP.

作者信息

Barsony J, Marx S J

机构信息

Mineral Metabolism Section, National Institute of Diabetes and Digestive and Kidney Diseases, Bethesda, MD 20892.

出版信息

Proc Natl Acad Sci U S A. 1990 Feb;87(3):1188-92. doi: 10.1073/pnas.87.3.1188.

DOI:10.1073/pnas.87.3.1188
PMID:2153973
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC53436/
Abstract

We developed a method for cAMP and cGMP immunocytology based upon fixation by microwave irradiation. Fixation by microwave irradiation prevented three problems found with other fixation methods: nucleotide loss from cells, nucleotide diffusion within cells, and chemical modification of immunologic epitopes. Six agonists (four that stimulate adenylate cyclase and two that stimulate guanylate cyclase) produced cAMP or cGMP accumulation patterns that were agonist-specific, dose-dependent, detectable at physiologic concentrations of hormone, and time-dependent within 15 sec to 30 min. cAMP accumulation after 1 mM forskolin was greatest in the nucleus. Isoproterenol, prostaglandin E2, or calcitonin caused initial accumulation of cAMP along the plasma membrane, but later accumulation was greater in the cytoplasm. With calcitonin the later accumulation of cAMP was selectively perinuclear and along the nuclear membrane. Sodium nitroprusside stimulated cGMP accumulation diffusely throughout the cytoplasm. Atrial natriuretic peptide initiated cGMP accumulation near the plasma membrane, and cGMP accumulation moved from there into the cytoplasm. In conclusion, microwave irradiation preserved cell structure and allowed visualization of expected as well as unsuspected changes in intracellular accumulation patterns of cAMP and cGMP.

摘要

我们开发了一种基于微波辐照固定的环磷酸腺苷（cAMP）和环磷酸鸟苷（cGMP）免疫细胞化学方法。微波辐照固定避免了其他固定方法中出现的三个问题：细胞内核苷酸丢失、细胞内核苷酸扩散以及免疫表位的化学修饰。六种激动剂（四种刺激腺苷酸环化酶，两种刺激鸟苷酸环化酶）产生的cAMP或cGMP积累模式具有激动剂特异性、剂量依赖性，在生理浓度的激素下可检测到，且在15秒至30分钟内呈时间依赖性。1 mM福司可林作用后，cAMP在细胞核中的积累量最大。异丙肾上腺素、前列腺素E2或降钙素导致cAMP最初沿质膜积累，但随后在细胞质中的积累量更大。对于降钙素，cAMP的后期积累选择性地位于核周并沿核膜分布。硝普钠刺激cGMP在整个细胞质中弥漫性积累。心房利钠肽在质膜附近引发cGMP积累，且cGMP积累从那里转移到细胞质中。总之，微波辐照保留了细胞结构，并使人们能够观察到cAMP和cGMP细胞内积累模式中预期以及未预料到的变化。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/e4de668b8cb7/pnas01028-0344-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/1946bbd4f9b6/pnas01028-0342-a.jpg
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https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/e4de668b8cb7/pnas01028-0344-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/1946bbd4f9b6/pnas01028-0342-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/311818854d3b/pnas01028-0342-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/deb4cdd145ca/pnas01028-0343-a.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/b6eec3274a96/pnas01028-0343-b.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/c01ab0585344/pnas01028-0343-c.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b153/53436/e4de668b8cb7/pnas01028-0344-a.jpg

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