Multiple time-gate module for fluorescence lifetime imaging

被引:77
作者
de Grauw, CJ [1 ]
Gerritsen, HC [1 ]
机构
[1] Univ Utrecht, Debye Inst, MBF, NL-3584 CC Utrecht, Netherlands
关键词
time gating; fluorescence; lifetime; two-photon excitation;
D O I
10.1366/0003702011952587
中图分类号
TH7 [仪器、仪表];
学科分类号
0804 ; 080401 ; 081102 ;
摘要
A versatile multiple time-gate fluorescence lifetime acquisition module is presented. The module is used as an add-on to a two-photon scanning laser microscope equipped with pulsed excitation for fluorescence lifetime imaging. It enables the recording of fluorescence lifetimes by capturing the intensity decay in four or eight time gates. The module allows fluorescence lifetime imaging at high count rates and with a high efficiency. The module is equipped with computer-controlled variable-delay lines for setting the gate widths. Decay times can be accurately measured in the 0.5 ns-1 mus range. Fluorescence lifetime imaging experiments on a number of single dye solutions with different (mono-exponential) fluorescence lifetimes are in excellent agreement with the reference values obtained with time-correlated single photon counting. Measurements on a test specimen exhibiting a bi-exponential decay demonstrate that the two decay components can be quantified with high accuracy. In a specimen containing six different lifetimes, in the range 2.8 to 12 ns, all lifetimes could be well distinguished. As an example, the results of a pH imaging experiment on oral biofilm using carboxyfluorescein are presented. Here, fluorescence lifetime imaging is used both to separate the short-lifetime autofluorescence from the carboxy-fluorescein fluorescence and to determine the pH in the biofilm.
引用
收藏
页码:670 / 678
页数:9
相关论文
共 38 条
  • [1] [Anonymous], 1991, TOPICS FLUORESCENCE
  • [2] AN ERROR ANALYSIS OF THE RAPID LIFETIME DETERMINATION METHOD FOR THE EVALUATION OF SINGLE EXPONENTIAL DECAYS
    BALLEW, RM
    DEMAS, JN
    [J]. ANALYTICAL CHEMISTRY, 1989, 61 (01) : 30 - 33
  • [3] Fluorescence lifetime imaging microscopy: spatial resolution of biochemical processes in the cell
    Bastiaens, PIH
    Squire, A
    [J]. TRENDS IN CELL BIOLOGY, 1999, 9 (02) : 48 - 52
  • [4] Microspectroscopic imaging tracks the intracellular processing of a signal transduction protein: Fluorescent-labeled protein kinase C beta I
    Bastiaens, PIH
    Jovin, TM
    [J]. PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1996, 93 (16) : 8407 - 8412
  • [5] A modified chemostat system to study the ecology of oral biofilms
    Bradshaw, DJ
    Marsh, PD
    Schilling, KM
    Cummins, D
    [J]. JOURNAL OF APPLIED BACTERIOLOGY, 1996, 80 (02): : 124 - 130
  • [6] FLUORESCENCE LIFETIME IMAGING USING A CONFOCAL LASER SCANNING MICROSCOPE
    BUURMAN, EP
    SANDERS, R
    DRAAIJER, A
    GERRITSEN, HC
    VANVEEN, JJF
    HOUPT, PM
    LEVINE, YK
    [J]. SCANNING, 1992, 14 (03) : 155 - 159
  • [7] THE USE OF EXOGENOUS FLUORESCENT-PROBES FOR TEMPERATURE-MEASUREMENTS IN SINGLE LIVING CELLS
    CHAPMAN, CF
    LIU, Y
    SONEK, GJ
    TROMBERG, BJ
    [J]. PHOTOCHEMISTRY AND PHOTOBIOLOGY, 1995, 62 (03) : 416 - 425
  • [8] Clegg R. M, 1996, Fluorescence Imaging Spectroscopy and Microscopy
  • [9] Imaging properties in two-photon excitation microscopy and effects of refractive-index mismatch in thick specimens
    de Grauw, GJ
    Vroom, JM
    van der Voort, HTM
    Gerritsen, HC
    [J]. APPLIED OPTICS, 1999, 38 (28) : 5995 - 6003
  • [10] Two-photon fluorescence lifetime imaging microscopy of macrophage-mediated antigen processing
    French, T
    So, PTC
    Weaver, DJ
    CoelhoSampaio, T
    Gratton, E
    Voss, EW
    Carrero, J
    [J]. JOURNAL OF MICROSCOPY-OXFORD, 1997, 185 : 339 - 353