Journal: Nature Communications

Article Title: Fluorescent sensors for imaging of interstitial calcium

doi: 10.1038/s41467-023-41928-w

Figure Lengend Snippet: Twelve combinations of export signal peptides and transmembrane/anchoring domains were used to target GreenT-EC to the surface of the cell. HeLa cells were transiently transfected with each construct and analyzed using flow cytometry. a Density plots of the negative control (non-transfected) indicating the gating strategy for selecting GreenT-EC positive single cells (indicated in slashed lines). Both the negative and the rest of the constructs displayed were resuspended in a MOPS buffer containing 3 mM CaCl 2 . b The fluorescence of GreenT-EC positive gated cells was further analyzed before and after the addition of EGTA (final concentration of 7.5 mM). Box-plots 90-10% are presented (whiskers are 90 and 10 percentiles, boxes are 75 and 25 percentiles and the center line correspond to the median). c Summary table compiling the main parameters of the experiment: Number of cells gated as GreenT-EC positive in each condition (n=cell count), mean fluorescence of GreenT-EC (mean fluoresce.), standard error of the mean (s.e.m) and the response calculated between the mean GreenT-EC fluorescence values before and after the addition of EGTA (ΔF/F 0 ). Source data are provided in the Source Data file.

Article Snippet: After centrifugation at 1000 g during 5 min, cells were resuspended in 1.5 ml MOPS buffer supplemented with 3 mM CaCl 2 / 1 mM MgCl 2 , and transferred to 5 ml round bottom tubes with cell strainer snap cap (Corning).

Techniques: Transfection, Construct, Flow Cytometry, Negative Control, Fluorescence, Concentration Assay, Cell Counting