title: Localizable and highly sensitive calcium indicator based on a BODIPY fluorophore.
abstract: We introduce here a new class of BODIPY-based Ca2+ indicators which can be derivatized with biological ligands that permit the localization of the indicators in living cells. The underivatized BODIPY-based Ca2+ indicator (BOCA-1) shows a 250-fold increase in fluorescence intensity upon Ca2+ binding. We also prepared its O6-benzylguanine (BG) derivative, BOCA-1-BG, which can be covalently and specifically linked to SNAP-tag fusion proteins in living cells. The indicator retains its properties as a highly sensitive Ca2+ indicator after conjugation to proteins, displaying a 180-fold increase in fluorescence intensity upon Ca2+ binding. We further demonstrated that BOCA-1-BG through reaction with localized SNAP-tag fusion proteins can be used to sense changes in Ca2+ concentrations in the nuclei and in the cytosol of live CHO-K1 cells. The high sensitivity of the indicator together with the possibility to selectively couple it to proteins of interest makes it a powerful tool for measuring local changes in Ca2+ concentrations in living cells.
authors.affiliation: Institute of Chemical Sciences and Engineering, Ecole Polytechnique Fédérale de Lausanne, CH 1015, Lausanne, Switzerland.
authors.forename: Mako
authors.initials: M
authors.lastname: Kamiya
authors.forename: Kai
authors.initials: K
authors.lastname: Johnsson
mesh_terms: D000818:Animals; D001896:Boron Compounds / Q000138:chemical synthesis / Q000737:chemistry*; D016466:CHO Cells; D002118:Calcium / Q000032:analysis*; D006224:Cricetinae; D003412:Cricetulus; D005456:Fluorescent Dyes / Q000737:chemistry*; D005998:Glycine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D006147:Guanine / Q000031:analogs & derivatives / Q000737:chemistry; D007202:Indicators and Reagents / Q000032:analysis; D008856:Microscopy, Fluorescence / Q000379:methods*; D011687:Purines / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D001896:Boron Compounds; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D011687:Purines; C064976:O(6)-benzylguanine; D006147:Guanine; D002118:Calcium; D005998:Glycine