Abstract

We describe imaging of calcium concentrations using the long-wavelength Ca2+ indicators, Calcium Green, Orange, and Crimson. The lifetimes of these probes were measured using the frequency-domain method and were found to increase from 50% to several fold in response to calcium. The two-dimensional images of the calcium concentration were obtained using a new apparatus for fluorescence lifetime imaging (FLIM). We also describe procedures to correct for the position-dependent frequency response of the gain-modulated image intensifier used in the FLIM apparatus. Importantly, the FLIM method does not require the probe to display shifts in the excitation or emission spectra. Using the FLIM method, calcium imaging is possible using probes which display changes in lifetime in response to calcium. Consequently, calcium imaging is possible with excitation wavelengths ranging from 488 to as long as 620 nm, where autofluorescence and/or photochemical damage is minimal. These probes are also suitable for calcium measurements of single cells using lifetime-based flow cytometry.?

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR008119-09
Application #
6444715
Study Section
Project Start
2001-04-01
Project End
2002-03-31
Budget Start
1997-10-01
Budget End
1998-09-30
Support Year
9
Fiscal Year
2001
Total Cost
$293,069
Indirect Cost

  Institution

Name
University of Maryland Baltimore
Department
Type
DUNS #
003255213
City
Baltimore
State
MD
Country
United States
Zip Code
21201

  Publications

Piao, Wenji; Shirey, Kari Ann; Ru, Lisa W et al. (2015) A Decoy Peptide that Disrupts TIRAP Recruitment to TLRs Is Protective in a Murine Model of Influenza. Cell Rep 11:1941-52
Hartman, Kira G; McKnight, Laura E; Liriano, Melissa A et al. (2013) The evolution of S100B inhibitors for the treatment of malignant melanoma. Future Med Chem 5:97-109
Fu, Yi; Zhang, Jian; Lakowicz, Joseph R (2009) Highly efficient detection of single fluorophores in blood serum samples with high autofluorescence. Photochem Photobiol 85:646-51
Zhang, Jian; Badugu, Ramachandram; Lakowicz, Joseph R (2008) Fluorescence Quenching of CdTe Nanocrystals by Bound Gold Nanoparticles in Aqueous Solution. Plasmonics 3:3-11
Zhang, Jian; Lakowicz, Joseph R (2007) Metal-enhanced fluorescence of an organic fluorophore using gold particles. Opt Express 15:2598-606
Fu, Yi; Lakowicz, Joseph R (2007) Single-Molecule Studies of Enhanced Fluorescence on Silver Island Films. Plasmonics 2:1-4
Ray, Krishanu; Szmacinski, Henryk; Enderlein, Joerg et al. (2007) Distance dependence of surface plasmon-coupled emission observed using Langmuir-Blodgett films. Appl Phys Lett 90:251116
Chowdhury, Mustafa H; Malyn, Stuart N; Aslan, Kadir et al. (2007) First Observation of Surface Plasmon-Coupled Chemiluminescence (SPCC). Chem Phys Lett 435:114-118
Zhang, Jian; Fu, Yi; Chowdhury, Mustafa H et al. (2007) Enhanced Forster Resonance Energy Transfer on Single Metal Particle. 2. Dependence on Donor-Acceptor Separation Distance, Particle Size, and Distance from Metal Surface. J Phys Chem C Nanomater Interfaces 111:11784-11792
Zhang, Jian; Fu, Yi; Lakowicz, Joseph R (2007) Luminescent images of single gold nanoparticles and their labeling on silica beads. Opt Express 15:13415-20

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