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A Photoconvertible Protein for Calcium Activity History Imaging
CaMPARI (Calcium Modulated Photoactivatable Ratiometric Integrator) is a photoconvertible protein construct that enables imaging of the integrated calcium activity of large populations of cells over defined time windows. This second generation of CaMPARI molecules has several advantages over their predecessors. By mutating the amino acids in the photophore domain, researchers improved green brightness by 50% and red brightness by 250%. Additionally, these new molecules have faster kinetics, and a lower photoconversion rate in low calcium conditions has resulted in a net 100-fold difference in green-to-red switching in low- vs. high-calcium conditions. These different CaMPARI proteins display a range of sensitivity in their calcium-binding, with Kd’s ranging from 100 nM to 1 uM of free calcium – an effective working sensitivity range spanning an order of magnitude.
Janelia researchers generated an Anti-CaMPARI-Red [4F6] antibody specific to the red form of the CaMPARI molecule. This allows for the system to be used in conjunction with IHC methods for fixed tissue analysis and facilitate the amplification of weaker red signals. The antibody is available from Ximbio and their distributors here.
DNA constructs for pAAV_hsyn_CaMPARI2, pAAV_hsyn_CaMPARI2_F391W, pAAV_hsyn_CaMPARI2_H396K, pAAV_hsyn_CaMPARI2_F391W-G395D, and pAAV_hsyn_CaMPARI2_L398T are available via Addgene.
AAV virus can be requested through Addgene's Viral Services.
Tg(elavl3:CaMPARI2)jf92 transgenic zebrafish are deposited to the ZIRC.
Drosophila expressing CaMPARI2 and CaMPARI2-L398T under UAS and LexA promoter in chromosome 2 (su(Hw)attP5), 3 (VK00005), or X (su(Hw)attP8) are available from the Bloomington Drosophila Stock Center (#78316-#78326).
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