HelloI am measuring the calcium concentration in cancer cells...
Hello
I am measuring the calcium concentration in cancer cells using Fura-2Am. I am unable to understand the Rmin and Rmax values needed to be taken for calibration of the experiment. I am using EGTA (3 uM) and Digitonin (100 um) for RMax and R min respectively. But I am not get Read more Read less

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Hi
And what about incubation time of EGTA or Cacl2
María Fernández-Velasco Author Answered Nov 5, 2014
Cell Communiation, Instituto de Investigaciones Biomédicas Alberto Sols, Spain
Hi
You should first incubated cells with FURA and then with EGTA.
Incubation with FURA is 30-40 min.
Hi
Thanks for the suggestion. Could you tell me what is the incubation time for Rmin and Rmax. Whether the cells should be simultaneously incubated with both EGTA (3 uM) and Fura-2am at the same time. Or first should i pre-incubate cell with EGTA followed by Fura2am loading.
María Fernández-Velasco Author Answered Nov 3, 2014
Cell Communiation, Instituto de Investigaciones Biomédicas Alberto Sols, Spain
Hello you should calculated R Min and Rmax separately
Hello
Thanks for your suggestion. But I just wanted to know that whether Rmin and Rmax will be calculated separately or in the same system
María Fernández-Velasco Author Answered Oct 30, 2014
Cell Communiation, Instituto de Investigaciones Biomédicas Alberto Sols, Spain
Hello,
maybe you can try to calibrate with 2.5 mM CaCl2 (Rmax) and 10 mM EGTA (Rmin). Morover there are some comercial kits to make a calibration curve (check in Invitrogen/Molecular Probes)
María/Lisardo
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