Get A Practical Guide to the Study of Calcium in Living Cells PDF

By Richard Nuccitelli, Leslie Wilson, Paul T. Matsudaira

ISBN-10: 0125228104

ISBN-13: 9780125228107

ISBN-10: 0125641419

ISBN-13: 9780125641418

A Practical consultant to the learn of Calcium in residing Cells describes renowned thoughts in addition to important do's and don't's and computing device courses. the quantity permits investigators to guage confocal photographs, use the most recent dyes, and layout Calcium buffers applicable to their learn wishes. This ebook is designed for laboratory use via graduate scholars, technicians, and researchers in lots of disciplines, starting from molecular to mobile degrees of research. Key beneficial properties * Describes concepts for detection of [Ca2+]I: Ca2+ - delicate microelectrodes * Fluorescent dyes * Luminescent proteins * comprises recommendations for perturbing intracellular Ca2+ * Covers targeted technique plus difficulties and pitfalls of every strategy * incorporates a sensible consultant to getting ready Ca2+ buffers with an easy-to-use desktop application * colour plates illustrate ideas equivalent to * Confocal ratio-imaging * Use of aequorin

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Additional info for A Practical Guide to the Study of Calcium in Living Cells

Example text

To increase the change in Ca" binding affinity on photolysis. nitr-8 was created with a 2-nitrobenzyl group on each aromatic ring of BAPTA. 026. Finally, nitr-9 is a dicarboxylate 2-nitrobenzhydrol with a low Ca2+ affinity that is unaffected by photolysis; this compound can be used to control for nonspecific effects of the photoproducts. Nitr-5 is the substance most often applied in biological experiments, largely because it was the first photolabile chelator to have most of the qualities of the ideal substance.

We often have been able to store Ca2+ calibration solutions for up to 2-3 months in polypropylene bottles (provided no organic substrate to foster bacterial growth is present). An accurate [Ca2+]standard is important for making Ca2+ buffers. Making accurate [Ca2+]using CaC12 2H20 typically used to make physiological solutions is difficult because the hydration state varies, making stoichiometric weighing imprecise. CaCO3 can be weighed more accurately, but has the disadvantage that the C02 must be driven off with prolonged heating and HCI, unless HC03 (which is a weak Ca2+buffer itself) is desired in the solutions.

Additionally, by adjusting the Ca2+loading or selecting chelator variants, the initial resting Ca2+ level could be set to somewhat higher or lower than the normal resting concentration. 3. The chelator should be chemically and photolytically stable. 4. Photolysis by a bright flash of light should allow rapid changes in the free Ca2+ level: this characteristic requires rapid photochemical and subsequent dark reactions of the chelator. 5 . Photolysis should be achievable with biologically appropriate wavelengths, which requires a high quantum efficiency and absorbance at wavelengths that readily penetrate cytoplasm but cause little biological damage, that is, that are not highly ionizing.

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A Practical Guide to the Study of Calcium in Living Cells by Richard Nuccitelli, Leslie Wilson, Paul T. Matsudaira


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