Picosecond wide-field time-correlated single photon counting fluorescence microscopy with a delay line anode detector

Journal Article · Mon Aug 15 00:00:00 EDT 2016 · Applied Physics Letters

DOI:https://doi.org/10.1063/1.4961054· OSTI ID:22590537

Hirvonen, Liisa M.; Le Marois, Alix; Suhling, Klaus ^[1]; Becker, Wolfgang; Smietana, Stefan ^[2]; Milnes, James; Conneely, Thomas ^[3]; Jagutzki, Ottmar ^[4]

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1. Department of Physics, King's College London, Strand, London WC2R 2LS (United Kingdom)
2. Becker & Hickl GmbH, Nahmitzer Damm 30, 12277 Berlin (Germany)
3. Photek Ltd., 26 Castleham Rd, Saint Leonards-on-Sea TN38 9NS (United Kingdom)
4. Institut für Kernphysik, Max-von-Laue-Str. 1, 60438 Frankfurt (Germany)

We perform wide-field time-correlated single photon counting-based fluorescence lifetime imaging (FLIM) with a crossed delay line anode image intensifier, where the pulse propagation time yields the photon position. This microchannel plate-based detector was read out with conventional fast timing electronics and mounted on a fluorescence microscope with total internal reflection (TIR) illumination. The picosecond time resolution of this detection system combines low illumination intensity of microwatts with wide-field data collection. This is ideal for fluorescence lifetime imaging of cell membranes using TIR. We show that fluorescence lifetime images of living HeLa cells stained with membrane dye di-4-ANEPPDHQ exhibit a reduced lifetime near the coverslip in TIR compared to epifluorescence FLIM.

• OSTI ID: 22590537
• Journal Information: Applied Physics Letters, Vol. 109, Issue 7; Other Information: (c) 2016 Author(s); Country of input: International Atomic Energy Agency (IAEA); ISSN 0003-6951
• Country of Publication: United States
• Language: English

Related Subjects

46 INSTRUMENTATION RELATED TO NUCLEAR SCIENCE AND TECHNOLOGY
71 CLASSICAL AND QUANTUM MECHANICS
GENERAL PHYSICS
ANODES
BIOMEDICAL RADIOGRAPHY
CELL MEMBRANES
FLUORESCENCE
HELA CELLS
ILLUMINANCE
IMAGE INTENSIFIERS
LIFETIME
MICROCHANNEL ELECTRON MULTIPLIERS
MICROSCOPES
MICROSCOPY
PHOTONS
READOUT SYSTEMS
REFLECTION
STAINS
TIME RESOLUTION