Measuring Intranuclear and Nuclear Envelope [Ca2+] vs. Cytosolic [Ca2+]
Nuclear Ca2+ regulates key cellular processes, including gene expression, apoptosis, assembly of the nuclear envelope, and nucleocytoplasmic transport. Quantification of subcellularly resolved Ca2+ signals is, therefore, essential for understanding physiological and pathological processes in various cell types. However, the properties of commonly used Ca2+-fluorescent indicators in intracellular compartments may differ, thus affecting the translation of Ca2+-dependent fluorescence changes into quantitative changes of Ca2+ concentration. Here, we describe technical approaches for reliable subcellular quantification of [Ca2+] in the cytoplasm vs. the nucleus and the nuclear envelope by in situ calibration of fluorescein-derived fluorescent indicators Fluo-4 and Fluo-5N.
Key wordsCa2+ concentration In situ calibration Nucleus Nuclear envelope Cytoplasm Fluo-4 Fluo-5N
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