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Personal profile

Research interests

Since joining King’s College London in February 2006, I have developed a programme of research spanning three separate areas of microscopy with the broad goal of investigating quantitative molecular imaging for the study of protein-protein interactions. My current research program focuses on the development of optical instruments to address fundamental biological questions regarding the dynamic interaction of protein partners within the cellular environment, furthering our understanding of cell signalling dynamics and control in cancer.

I have been involved in four high-profile collaborative imaging projects since joining KCL funded by EPSRC, Cancer Research UK and BBSRC (2 projects) respectively the details of which are summarised below.

The nature of my work is, by definition, multidisciplinary and I have been successful in both acquiring funding in this area and prosecuting the work throughout my career.

Optical proteomics technology: High content screening of protein-protein interactions: I have been involved in an initiative to develop ‘optical proteomic technology for in situ analysis of protein interaction networks’ since its inception prior to joining the college when I collaborated with Prof T Ng for a number of years in developing time-resolved multiphoton microscopy for FRET applications. This collaboration, involving a number of research groups within the college and led by Prof Malcolm Irving (KCL), aimed to develop high-throughput/content optical screening approaches for cell based assays of protein-protein interactions. Such a development of high-throughput screening (HTS) for protein-protein and protein-effecter interactions represents a paradigm shift in proteomics technology. For my component of the project, a fluorescence anisotropy read out was initially considered for homo-FRET experiments of protein dimerisation but recent work by Piston and co-workers  had indicated measurement of acceptor anisotropy in fluorescent protein FRET experiments could provide a sensitive read-out for FRET efficiency. I developed a simplified, open architecture, microscope platform with both wide-field steady-state anisotropy (using a QuadView Image Splitter) and laser scanning TCSPC fluorescence lifetime imaging modes. The rather surprising outcome of this work is that the acceptor anisotropy methodology compares very favourably with the more established donor FLIM methods that we had been using to date. Specifically, in measurements of a Cdc42-Pak1 Riachu biosensor in Jurkat T-Cells we were able to undertake measurements of the sensor at 1 fps which otherwise would have taken minutes by FLIM but with a degree of quantification of FRET that is comparable. Furthermore, the dynamic range of the technique is significantly greater for acceptor anisotropy, since we are sensitive predominantly to molecules undergoing FRET seeing a change of ~0.2 in anisotropy compared with a 200 ps change in the fluorescence lifetime for 10% FRET efficiency. We have recently shown that measurements of FRET by FLIM and anisotropy are correlated in high-content screens of inhibitor and siRNA  libraries. The great advantage of the acceptor anisotropy method is that it is much faster; a 96 well plate taking just 30 mins by acceptor anisotropy compared to 10 hours using donor FLIM. The most recent work characterising HCS and making a direct comparison between donor FLIM and acceptor anisotropy has been published in the open access journal PLoS ONE.

For proteomic screens of protein-protein interactions envisaged by the optical proteomics programme, a need for high-throughput screening of (potentially) millions of constructs is very clear. Without a significant advance in either parallelisation (improvement in widefield techniques to provide the required temporal resolution)  or counting rate for time-correlated single photon counting technologies (currently limited to ~5 million events per second dependent on hardware and excitation rate) it is clear that laser scanning microscopy will not fulfil this need for adherent cell assays. Conversely, HTP screening using flow cytometry techniques is very much more tractable. My group has developed a microfluidics based flow cytometer for FRET based screening applications. The concept of the system is very simple and relies on the flow of cells though a focused laser beam. The fluorescence excited in the cells containing fluorescent proteins or labelled with antibodies is detected using the burst integrated fluorescence lifetime technique. To date, we have measured cell populations with four biosensor constructs as a proof of principle and mixtures of a biosensors enabling us to test analysis routines and sorting concepts. Initial work with an EGF phosphorylation assay is encouraging.

Development of Multiphoton Microscopy: Throughout my research career I have been involved in the development of Multiphoton fluorescence lifetime imaging. When I arrived at KCL, I was able to quickly construct a custom multiphoton FLIM system with funds partially provided by the department, the Royal Society and Guy’s & St Thomas’ Cancer Trust. The system was designed to enable us to investigate a novel evanescent wave fluorescence lifetime imaging technique for protein-interaction monitoring at the cell membrane. Initial studies by a joint PhD student (Dr Quirke) have shown that this is possible although there are a number of limitations to the technique.

Following our success in obtaining research funding for the Cancer Research UK, Comprehensive Cancer Imaging Centre I have instigated a programme of work to investigate adaptive optics (AO) in multiphoton microscopy. My group is developing a multiphoton FLIM microscope specifically for in vivo studies in rodents and will examine both feedback controlled AO optimisation and a sensor based method for mapping the optical aberrations. A collaboration with Frederick Geissman (Centre for Molecular & Cellular Biology of Inflammation) has led us to develop a third multiphoton FLIM instrument within Prof Geissman’s lab which is based on our flexible design and includes adaptive optics elements developed using a spatial light modulator.

The most exciting development in my lab is currently the application of multi-beam multiphoton microscopy to parallelise fluorescence lifetime imaging using a newly developed CMOS SPAD camera system which incorporates pixel-by-pixel 50 ps timing for TCSPC. With Prof R. Henderson (Edinburgh) and Dr K. Suhling (Physics, KCL), I am developing a program of work to apply a novel camera system to provide fast frame rate FLIM data for multiphoton microscopy. We were successful in obtaining funding from BBSRC for the work in 2011 and work began in November. At current data rates, the imaging speed-up is modest (factor of 10 over current methodologies) but with firmware and hardware modifications this is expected to be improved to a factor 100. Publications  and conference presentations were presented at Photonics West 2013 in February. Extension of these and additional techniques has been proposed as part of the recent successful application to MRC for the Next Generation Optical Microscopy Initiative.

Single molecule imaging and spectroscopy: In order to further our understanding of single-molecule spectroscopy, my group embarked on an exploration of single-molecule fluorescence lifetime spectroscopy based on TCSPC and the burst-integrated fluorescence lifetime technique (BiFL). With a PhD student (Dr Pathmananthan), a TCSPC-BiFL system was developed and tested using a number of fluorophores including quantum dots and fluorescent proteins. The binding of biotin labelled acceptors to a streptavidin coated quantum dot was investigated and shown to be an interesting model system for protein hetero dimerisation – since Steptavidin may bind to upto 4 acceptors. This work led to collaboration with Dr Paul Barber (Oxford and Randall Division) and Prof Ton Coolen (Mathematics and Randall Division) to develop Bayesian analysis tools for single molecule burst detection and fluorescence lifetime analysis.

Having obtained funding from BBSRC (In collaboration with Peter Parker, Tony Ng (Cancer Division), Ton Coolen (Mathematics) et al.) and industry (UCB, in collaboration with Andrew Beavil, Randall Division) to pursue single-molecule imaging, I have developed two TIRF imaging systems based on Nikon platforms with custom modifications for laser excitation and software. Both systems may be used for either ensemble measurements of cell membrane receptors or single-molecule imaging studies of protein-protein interaction. Following recent advances in the field of super-resolution microscopy we have added capability to undertake STORM and PALM experiments in two colours using an image splitter. I am currently investigating methods to exploit FRET in a single-molecule imaging context.

Research interests (short)

Protein-protein interactions; Multiphoton Fluorescence Lifetime Imaging; Single molecule imaging; FRET; Fluorescence anisotropy

Click here for the Ameer-Beg group webpage

Expertise related to UN Sustainable Development Goals

In 2015, UN member states agreed to 17 global Sustainable Development Goals (SDGs) to end poverty, protect the planet and ensure prosperity for all. This person’s work contributes towards the following SDG(s):

  • SDG 3 - Good Health and Well-being

Education/Academic qualification

Doctor of Philosophy, Femtosecond laser interactions in the condensed phase: Application to transient absorption and materials processing, University of Central Lancashire

Award Date: 1 Jan 1999

Master of Science, University of Essex

Award Date: 1 Jan 1994

Bachelor of Science, University of Essex

Award Date: 1 Jan 1993


  • QC Physics
  • Multiphoton microscopy
  • FRET
  • Fluorescience Lifetime Imaging
  • High Content Screening
  • Fluorescence Anisotropy
  • Super-resolution Imaging
  • Single molecule imaging
  • TIRF


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