Multifocal multiphoton microscopy based on multianode photomultiplier tubes

被引:89
作者
Kim, Ki Hean
Buehler, Christof
Bahlmann, Karsten
Ragan, Timothy
Lee, Wei-Chung A.
Nedivi, Elly
Heffer, Erica L.
Fantini, Sergio
So, Peter T. C. [1 ]
机构
[1] MIT, Dept Mech Engn, Cambridge, MA 02139 USA
[2] Novartis Inst Biomed Res, A-1235 Vienna, Austria
[3] TissueVis Inc, Somerville, MA 02143 USA
[4] MIT, Dept Brain & Cognit Sci, Cambridge, MA 02139 USA
[5] MIT, Dept Biol, Cambridge, MA 02139 USA
[6] Tufts Univ, Dept Biomed Engn, Medford, MA 02155 USA
[7] MIT, Biol Engn Div, Cambridge, MA 02139 USA
来源
OPTICS EXPRESS | 2007年 / 15卷 / 18期
关键词
D O I
10.1364/OE.15.011658
中图分类号
O43 [光学];
学科分类号
070207 ; 0803 ;
摘要
Multifocal multiphoton microscopy (MMM) enhances imaging speed by parallelization. It is not well understood why the imaging depth of MMM is significantly shorter than conventional single-focus multiphoton microscopy (SMM). In this report, we show that the need for spatially resolved detectors in MMM results in a system that is more sensitive to the scattering of emission photons with reduced imaging depth. For imaging depths down to twice the scattering mean free path length of emission photons (2 x l(s)(em)), the emission point spread function (PSFem) is found to consist of a narrow, diffraction limited distribution from ballistic emission photons and a broad, relatively low amplitude distribution from scattered photons. Since the scattered photon distribution is approximately 100 times wider than that of the unscattered photons at 2 x l(s)(em), image contrast and depth are degraded without compromising resolution. To overcome the imaging depth limitation of MMM, we present a new design that replaces CCD cameras with multi-anode photomultiplier tubes (MAPMTs) allowing more efficient collection of scattered emission photons. We demonstrate that MAPMT-based MMM has imaging depth comparable to SMM with equivalent sensitivity by imaging tissue phantoms, ex vivo human skin specimens based on endogenous fluorophores, and green fluorescent protein (GFP) expressing neurons in mouse brain slices. (c) 2007 Optical Society of America.
引用
收藏
页码:11658 / 11678
页数:21
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