A custom-made two-photon microscope and deconvolution system

被引:137
作者
Majewska, A [1 ]
Yiu, G [1 ]
Yuste, R [1 ]
机构
[1] Columbia Univ, Dept Biol Sci, New York, NY 10027 USA
来源
PFLUGERS ARCHIV-EUROPEAN JOURNAL OF PHYSIOLOGY | 2000年 / 441卷 / 2-3期
关键词
fluoview; laser; pockels; scanning; spines; XCOSM;
D O I
10.1007/s004240000435
中图分类号
Q4 [生理学];
学科分类号
071003 [生理学];
摘要
We describe in detail a custom-built two-photon microscope based on a modified confocal scanhead (Olympus Fluoview) and mode-locked Ti:sapphire laser (Coherent Mira 900). This system has internal detectors as well as external whole-field detection and an electrooptical modulator for blanking the beam on flyback and effecting fast changes in excitation intensity. This microscope can be used in deep, scattering samples for quantitative measurements with a wide range of fluorophores (GFP, fura, calcium green, calcium orange, fluo-3, DiI, DiO, fluorescein, rhodamine), for fluorescent photobleaching recovery and for uncaging. Images obtained with this system can be deconvolved with the Estimation Maximization algorithm using the program XCOSM (freeware available at: http://www.ibc.wustl.edu/bcV xcosm/).
引用
收藏
页码:398 / 408
页数:11
相关论文
共 23 条
[1]
Brown E B, 1998, Methods Enzymol, V291, P356
[2]
Measurement of molecular diffusion in solution by multiphoton fluorescence photobleaching recovery [J].
Brown, EB ;
Wu, ES ;
Zipfel, W ;
Webb, WW .
BIOPHYSICAL JOURNAL, 1999, 77 (05) :2837-2849
[3]
Photolysis of caged calcium in femtoliter volumes using two-photon excitation [J].
Brown, EB ;
Shear, JB ;
Adams, SR ;
Tsien, RY ;
Webb, WW .
BIOPHYSICAL JOURNAL, 1999, 76 (01) :489-499
[4]
CAJAL SRY, 1891, REV CIENCIAS MED BAR, V22, P23
[5]
ENHANCED 3-DIMENSIONAL RECONSTRUCTION FROM CONFOCAL SCANNING MICROSCOPE IMAGES .2. DEPTH DISCRIMINATION VERSUS SIGNAL-TO-NOISE RATIO IN PARTIALLY CONFOCAL IMAGES [J].
CONCHELLO, JA ;
KIM, JJ ;
HANSEN, EW .
APPLIED OPTICS, 1994, 33 (17) :3740-3750
[7]
2-PHOTON LASER SCANNING FLUORESCENCE MICROSCOPY [J].
DENK, W ;
STRICKLER, JH ;
WEBB, WW .
SCIENCE, 1990, 248 (4951) :73-76
[8]
Diaspro A, 1999, MICROSC RES TECHNIQ, V47, P196, DOI 10.1002/(SICI)1097-0029(19991101)47:3<196::AID-JEMT5>3.0.CO
[9]
2-V
[10]
Developmental regulation of spine motility in the mammalian central nervous system [J].
Dunaevsky, A ;
Tashiro, A ;
Majewska, A ;
Mason, C ;
Yuste, R .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1999, 96 (23) :13438-13443