Graphically represented calcium oscillations are continuous up and down peaks. They are more compact in cases of cell migration and cancer experiments. How can this data be analyzed with perfect prediction and be modeled? The data usually looks like the following figure.
Hi Sarpras,
I almost everyday measure cellular calcium in Fura-2 loaded cells and analyze the calcium oscillation data. The way I do it, I am trying to summarize:
1. Depending upon the imaging system you are working with you may need to post- or pre-calibrate each experiment for fluorescent ratios to actual calcium concentrations. For post-calibration at the end of individual experiment cells have to be treated with 5 μM ionomycin (Calbiochem, San Diego, CA) to permit calcium equilibration with the external solution. The superfusate then needs to be switched to a calcium-free solution containing 50 mM EGTA in the continued presence of ionomycin to obtain the minimum ratio. The minimal and maximal values were used to calculate [Ca2+]i following the formula provided by the manufacturer of the dye (Hartford AK et al-Glia. 2004 Feb;45(3):229-37)
2. For pre-calibration purpose the relationship between fluorescent ratios and calcium is usually established using a fura-2 calcium calibration kit (Life Technologies, Cat#. C-3008MP). In brief, fluorescence ratios (340 nm/380 nm) are derived using a set of standards containing 0-, 38-, 100-, 225-, 351-, 602-, and 1350-nM concentrations of free calcium. A calibration curve is then constructed by plotting the fura-2 fluorescence ratios (F340 nm/F380 nm) against the free calcium concentrations (Mandal A et al-Invest Ophthalmol Vis Sci. 2010 Jun; 51(6):3129-38).
2. In each experiment, data from 15 to 30 individual cells on the coverslip/dish are averaged and considered as n =1. You can take help of MS Excel to analyze the data.
We record calcium data for at least 5-10 individual experiments and each experiment consists 15 to 30 individual cells.
I am sharing these two papers as a combined pdf file as a ready reference.
Good luck!!
We are modeling ca signaling, please check our work: https://www.researchgate.net/publication/49835443_Inositol_(145)-trisphosphate_receptor_microarchitecture_shapes_Ca2_puff_kinetics?ev=prf_pub
and
https://www.researchgate.net/publication/7979813_Modeling_stochastic_Ca2_release_from_a_cluster_of_IP3-sensitive_receptors?ev=prf_pub
Article Inositol (1,4,5)-Trisphosphate Receptor Microarchitecture Sh...
Article Modeling stochastic Ca2+ release from a cluster of IP3-sensi...
Lusi Diambra: Thank you for this help. These paper are very much helpful for me
I have a question in relation to Amritlal Mandal's informative answer. I presume after you've carried out your 5-10 individual experiments, for each of them you will calculate a mean value and standard deviation of the signal from however many cells were examined in each experiment. After that you probably calculate a mean value of the mean signals from the 5-10 experiments. Assuming the above is correct, what quantity do you report as error on the mean value of your set of experiments? Normally, in papers of this sort one reads, for example, 'data from n=12 experiments are presented as mean +/- SEM'. What I wonder is whether that really is the SE of the mean of means; or should it perhaps be a pooled standard deviation?
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