RT info:eu-repo/semantics/article
T1 Measuring [Ca2+] in the endoplasmic reticulum with aequorin
A1 Álvarez Martín, Javier
A1 Montero Zoccola, María Teresa
K1 Calcio - Metabolismo
AB The photoprotein aequorin was the first probe used to measure specifically the [Ca2+] inside the lumen ofthe endoplasmic reticulum ([Ca2+]ER) of intact cells and it provides values for the steady-state [Ca2+]ER, around 500 M,that closely match those obtained now by other procedures. Aequorin-based methods to measure [Ca2+]ER offer severaladvantages: (i) targeting of the probe is extremely precise; (ii) the use of low Ca2+-affinity aequorin allows covering alarge dynamic range of [Ca2+], from 10−5 to 10−3 M; (iii) aequorin is nearly insensitive to changes in Mg2+ or pH, hasa high signal-to-noise ratio and calibration of the results in [Ca2+] is made straightforward using a simple algorithm;and (iv) the equipment required for luminescence measurements in cell populations is simple and low-cost. On thenegative side, this technique has also some disadvantages: (i) the relatively low amount of emitted light makes difficultperforming single-cell imaging studies; (ii) reconstitution of aequorin with coelenterazine requires previous completedepletion of Ca2+ of the ER for 1–2 h, a maneuver that may result in deleterious effects in some cells; (iii) becauseof the high rate of aequorin consumption at steady-state [Ca2+]ER, only relatively brief experiments can be performed;and (iv) expression of ER-targeted aequorin requires previous transfection or infection to introduce the appropriate DNAconstruct, or alternatively the use of stable cell clones. Choosing aequorin or other techniques to measure [Ca2+]ER willdepend of the correct balance between these properties in a particular problem.
PB Elsevier Ltd.
SN 0143-4160
YR 2002
FD 2002
LK http://uvadoc.uva.es/handle/10324/5957
UL http://uvadoc.uva.es/handle/10324/5957
LA eng
NO Cell Calcium, 2002, vol. 32, n. 5-6, p. 251-260
NO Producción Científica
DS UVaDOC
RD 22-nov-2024