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CalQuo: Automated, Simultaneous Single-Cell And Population-Level Quantification Of Intracellular Ca 2+ Responses

Detecting intracellular calcium signaling with fluorescent calcium indicator dyes is often coupled with microscopy techniques to follow the activation state of non-excitable cells, including lymphocytes. However, the analysis of global intracellular calcium responses both at the single-cell level and in large ensembles simultaneously has yet to be automated. We developed a new software package, CalQuo (Calcium Quantification), which allows the automated analysis and simultaneous monitoring of global fluorescent calcium reporter-based signaling responses in up to 1000 single cells per experiment, at temporal resolutions of sub-seconds to seconds. CalQuo quantifies the number and fraction of responding cells, the temporal dependence of calcium signaling and provides global and individual calcium-reporter fluorescence intensity profiles. 

CalQuo is a MATLAB based software package developed at the University of Oxford. CalQuo is copyright protected and licensed with Isis Innovation (www.isis-innovation.com/). CalQuo  allows   the   automated   analysis   and simultaneous monitoring of fluorescent calcium reporter­ based signaling responses in up to 1000 single cells per experiment. As output, CalQuo delivers the number and fraction of responding cells, their individual and characteristic calcium­ reporter fluorescence intensity profiles, and the high­resolution temporal dependence of calcium signaling. CalQuo is well suited to studying other cell signaling processes, such as the release of proteins orother molecules, if these molecules can be detected using labels.

Further information:

Fritzsche M et al, CalQuo: automated, simultaneous single-cell and population-level quantification of intracellular Ca 2+ responses., Scientific Reports, 2015.

 

CalQuo2: Automated Fourier-Space, Population-Level Quantification Of Global Intracellular Calcium Responses

To expand upon the capabilities of CalQuo to recognize and quantify calcium responses in individual cells within a sample up to 1000 cells, we have developed an updated version, CalQuo2. Using the Fourier transform to convert calcium responses into the frequency space, CalQuo2 now requires only one (instead of five) user-defined parameter in determining a triggering from a non-triggering cell. Additionally, CalQuo2 can determine the fraction of cells that have a single peaked compared to the fraction of cells that have multi-peak calcium fluxes.  

CalQuo2 is a MATLAB based software package developed at the University of Oxford. CalQuo is copyright protected and licensed with Isis Innovation. The new version employs Fourier-space analysis algorithm and is currently under review in a peer-reviewed journal.

Contact: 

Please contact for further information: marco.fritzsche (at) rdm.ox.ac.uk.

 
Figure taken from Fritzsche et al, Scientific Reports, 2015.  Representative intensity profiles  R ( t ) (upper panel) and their derivatives d R ( t )/d t  (lower panel); average over 200-650 individual T-cells (black circles: raw data, mangenta line: Savitzky–Golay interpolation for signaling T-cells, purple line: data of non-signaling cells).  CalQuo determines the characteristic times for the landing (grey shaded area) and signaling event (magenta shaded area) from the characteristic peaks in d R ( t )/d t . Error bars representing s.d.m.

Figure taken from Fritzsche et al, Scientific Reports, 2015.

Representative intensity profiles R(t) (upper panel) and their derivatives dR(t)/dt (lower panel); average over 200-650 individual T-cells (black circles: raw data, mangenta line: Savitzky–Golay interpolation for signaling T-cells, purple line: data of non-signaling cells). CalQuodetermines the characteristic times for the landing (grey shaded area) and signaling event (magenta shaded area) from the characteristic peaks in dR(t)/dt. Error bars representing s.d.m.

 

Contact: 

Please contact for further information: marco.fritzsche (at) rdm.ox.ac.uk or  ricardfe (at) stanford.edu.

Download:

CalQuo_ver01.tar.gz

 

 
Figure taken from Lee AM & Colin-York et al, Scientific Reports 2017.  Representative normalized Ca2+ based fluorescence intensity plot of a Jurkat T-cell upon landing on a 4D:MHC I functionalized coverslip with a subset of cells. Scale bar is 100 µm.

Figure taken from Lee AM & Colin-York et al, Scientific Reports 2017.

Representative normalized Ca2+ based fluorescence intensity plot of a Jurkat T-cell upon landing on a 4D:MHC I functionalized coverslip with a subset of cells. Scale bar is 100 µm.

Further information:

Lee AM, Colin-York H et al, CalQuo2 : Automated Fourier-space, population-level quantification of global intracellular calcium responses, Scientific Reports, 2017.

Download:

CalQuo2_ver01.tar.gz

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