Complementarity of PALM and SOFI for super-resolution live-cell imaging of focal adhesions

被引：66

作者：

Deschout, Hendrik ^{[1
,2
]}

Lukes, Tomas ^{[1
,2
,3
]}

Sharipov, Azat ^{[1
,2
]}

Szlag, Daniel ^{[1
,2
]}

Feletti, Lely ^{[1
,2
]}

Vandenberg, Wim ^{[4
]}

Dedecker, Peter ^{[4
]}

Hofkens, Johan ^{[4
]}

Leutenegger, Marcel ^{[5
]}

Lasser, Theo ^{[1
,2
]}

Radenovic, Aleksandra ^{[1
,2
]}

机构：

[1] Ecole Polytech Fed Lausanne, Lab Nanoscale Biol, STI IBI, Stn 17, CH-1015 Lausanne, Switzerland

[2] Ecole Polytech Fed Lausanne, STI IBI, Lab Opt Biomed, Stn 17, CH-1015 Lausanne, Switzerland

[3] Czech Tech Univ, FEE, Dept Radioelect, Tech 2, Prague 16627 2, Czech Republic

[4] Univ Leuven, Dept Chem, Celestijnenlaan 200F, B-3001 Heverlee, Belgium

[5] Max Planck Inst Biophys Chem, Abt NanoBiophoton, Fassberg 11, D-37077 Gottingen, Germany

来源：

NATURE COMMUNICATIONS | 2016年 / 7卷

基金：

欧盟地平线“2020”; 瑞士国家科学基金会;

关键词：

PHOTOACTIVATED LOCALIZATION MICROSCOPY; RESOLUTION; PROBES; NANOSCOPY; DYNAMICS;

D O I：

10.1038/ncomms13693

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

Live-cell imaging of focal adhesions requires a sufficiently high temporal resolution, which remains a challenge for super-resolution microscopy. Here we address this important issue by combining photoactivated localization microscopy (PALM) with super-resolution optical fluctuation imaging (SOFI). Using simulations and fixed-cell focal adhesion images, we investigate the complementarity between PALM and SOFI in terms of spatial and temporal resolution. This PALM-SOFI framework is used to image focal adhesions in living cells, while obtaining a temporal resolution below 10 s. We visualize the dynamics of focal adhesions, and reveal local mean velocities around 190 nmmin (-1). The complementarity of PALM and SOFI is assessed in detail with a methodology that integrates a resolution and signal-to-noise metric. This PALM and SOFI concept provides an enlarged quantitative imaging framework, allowing unprecedented functional exploration of focal adhesions through the estimation of molecular parameters such as fluorophore densities and photoactivation or photoswitching kinetics.

引用

页数：11

共 37 条

[1] Quantitative Photo Activated Localization Microscopy: Unraveling the Effects of Photoblinking [J].

Annibale, Paolo ;

Vanni, Stefano ;

Scarselli, Marco ;

Rothlisberger, Ursula ;

Radenovic, Aleksandra .

PLOS ONE, 2011, 6 (07)

[2] Identification of clustering artifacts in photoactivated localization microscopy [J].

Annibale, Paolo ;

Vanni, Stefano ;

Scarselli, Marco ;

Rothlisberger, Ursula ;

Radenovic, Aleksandra .

NATURE METHODS, 2011, 8 (07) :527-528

[3]

Annibale Paolo., 2012, OPTICAL NANOSCOPY, V1, P1, DOI DOI 10.1186/2192-2853-1-9

[4] Fourier ring correlation as a resolution criterion for super-resolution microscopy [J].

Banterle, Niccolo ;

Khanh Huy Bui ;

Lemke, Edward A. ;

Beck, Martin .

JOURNAL OF STRUCTURAL BIOLOGY, 2013, 183 (03) :363-367

[5] Imaging intracellular fluorescent proteins at nanometer resolution [J].

Betzig, Eric ;

Patterson, George H. ;

Sougrat, Rachid ;

Lindwasser, O. Wolf ;

Olenych, Scott ;

Bonifacino, Juan S. ;

Davidson, Michael W. ;

Lippincott-Schwartz, Jennifer ;

Hess, Harald F. .

SCIENCE, 2006, 313 (5793) :1642-1645

[6] Live Intracellular Super-Resolution Imaging Using Site-Specific Stains [J].

Carlini, Lina ;

Manley, Suliana .

ACS CHEMICAL BIOLOGY, 2013, 8 (12) :2643-2648

[7]

Cox S, 2012, NAT METHODS, V9, P195, DOI [10.1038/nmeth.1812, 10.1038/NMETH.1812]

[8] Assessing resolution in super-resolution imaging [J].

Demmerle, Justin ;

Wegel, Eva ;

Schermelleh, Lothar ;

Dobbie, Ian M. .

METHODS, 2015, 88 :3-10

[9] Fast, background-free, 3D super-resolution optical fluctuation imaging (SOFI) [J].

Dertinger, T. ;

Colyer, R. ;

Iyer, G. ;

Weiss, S. ;

Enderlein, J. .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2009, 106 (52) :22287-22292

[10]

Durisic N, 2014, NAT METHODS, V11, P156, DOI [10.1038/NMETH.2784, 10.1038/nmeth.2784]

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