We have worked since 1997 to develop practical technology for rapid time-gated FLIM utilising gated optical image intensifier technology, which we have utilised in microscopes, endoscopes, multiwell plate readers and tomographic imaging instrumentation. We gratefully acknowledge support from BBSRC, EPSRC, MRC, Imperial College London, the Wellcome Trust and the DTI/TSB, as well as advice and collaborations concerning the development and applications of FLIM instrumentation with AstraZeneca, Fianium Ltd, GE Healthcare, GSK, Kentech Instruments Ltd, PerkinElmer and Pfizer.

OPS

Thanks to support from an EPSRC IAA award, we are able to present here the information and software tools to enable research groups to replicate our high content analysis (HCA) instrumentation utilising time-gated fluorescence lifetime imaging (FLIM) for non-commercial use.

This software and descriptions of instrumentation are being made available in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public License for more details concerning the software.

The key components required to construct and apply our openFLIM-HCA platform are indicated in the diagram below and software tools can be reached from the links below.

OpenFLIM-HCA

Acquisition software

This software controls the data acquisition and is written in MicroManager. It can be configured to upload the locally saved data to an OMERO server. Please click here to access the software. For instructions on how to use the software, please access our wiki here.

OPH

Data management software

FLIM-HCA data can be uploaded to an OMERO 5.0 server from where it can be viewed, downloaded and shared. For more information, please go to the Open Microscopy Environment (OME) website.

To see an example of shared multiwell plate FLIM data used to support a journal publication, please click here.

FLIM data analysis software

We undertake FLIM data analysis using FLIMfit, an open source software tool and an OMERO client that is designed to facilitate analysis and visualisation of time-resolved FLIM data including from experiments utilising time-correlated single photon counting (TCSPC) and wide-field time-gated imaging.  FLIMfit provides tools for fitting to monoexponential and more complex data models on a per pixel basis as well as offering global binning (averaging over user-defined regions of interest) and global fitting (simultaneously fitting all pixels over one or more fields of view under the assumption that component lifetimes are spatially invariant). It can incorporate image segmentation and utilises a highly efficient algorithm that can globally fit FLIM data to complex decay models with modest photon numbers, across hundreds of fields of view, requiring only tens of seconds of processing time on a reasonable desktop computer. This makes it useful to analyse time series or multiwell plate FLIM data. Please see Warren et al, 2013 for further details.

FLIMfit is continuously being updated and may be downloaded from here.

For information concerning the operation of our FLIM instrumentation, please g o to http://docs.flimfit.org/

Selection of publications concerning FLIM-HCA

High speed unsupervised fluorescence lifetime imaging confocal multiwell plate reader for high content analysis
C. B. Talbot, J. McGinty, D. M. Grant, E. J. McGhee, D. M. Owen, W. Zhang, T. D. Bunney, I. Munro, B. Isherwood, R. Eagle, A. Hargreaves, M. Katan, C. Dunsby, M. A. A. Neil and P.M. W. French
J. Biophotonics 1 (2008) 514–521, DOI :10.1002/jbio.200810054

FLIM FRET technology for drug discovery: automated multiwell plate high content analysis, multiplexed readouts and application in situ,
S. Kumar, D. Alibhai, A. Margineanu, R. Laine, G. Kennedy, J. McGinty, S. Warren, D. Kelly, Y. Alexandrov, I. Munro, C. Talbot, D. W. Stuckey, C. Kimberly, B. Viellerobe, F. Lacombe, E. W.-F. Lam, H. Taylor, M. J. Dallman, G. Stamp, E. J. Murray, F. Stuhmeier, A. Sardini, M. Katan, D. S. Elson, M. A. A. Neil, C. Dunsby and P. M. W. French
ChemPhysChem 12 ( 2011), 627-633, DOI: 10.1002/cphc.201000874

Automated fluorescence lifetime imaging plate reader and its application to Förster resonant energy transfer readout of Gag protein aggregation
D. Alibhai, D. J. Kelly, S. Warren, S. Kumar, A. Margineanu, R. A. Serwa, E. Thinon, Y. Alexandrov, E. J. Murray, F. Stuhmeier, E. W. Tate, M. A.A. Neil, C. Dunsby and P. M.W. French,
J. Biophotonics 6 (2012) 398-408, DOI: 10.1002/jbio.201200185.

S.C. Warren, A. Margineanu, D. Alibhai, D.J. Kelly, C. Talbot, Y. Alexandrov, I. Munro, M. Katan, C. Dunsby and P.M.W. French, Rapid global fitting of large fluorescence lifetime imaging microscopy datasets,
PLoS ONE 8(2013) e70687; DOI: 10.1371/journal.pone.0070687

An automated multiwell plate reading FLIM microscope for live cell autofluorescence lifetime assays
D. J. Kelly, S. C. Warren, S. Kumar, J. L. Lagarto, B T. Dyer, A. Margineanu1, E. W.-F. Lam, C. Dunsby and P. M. W. French
J. Innov. Opt. Health Sci.. 7 (2014) 1450025-15 pages,  DOI: 10.1142/S1793545814500254

Automated multiwell fluorescence lifetime imaging for Förster resonant energy transfer assays and High Content Analysis,
D. J. Kelly, S. C. Warren, D. Alibhai, S. Kumar, Y. Alexandrov, I. Munro, A. Margineanu, J. McCormack, N. J. Welsh, R. A. Serwa, E. Thinon, M. Kongsema, J. McGinty, C. Talbot, E. J. Murray, F. Stuhmeier, M. A.A. Neil, E. W. Tate, V. M. M. Braga, E. W.-F. Lam, C. Dunsby and P. M.W. French
Analytical Methods, 7 (2015), 4071-4089, DOI: 10.1039/C5AY00244C