Improved localization of cellular membrane receptors using combined fluorescence microscopy and simultaneous topography and recognition imaging

被引：35

作者：

Duman, M.

Pfleger, M.

Zhu, R. ^{[4
]}

Rankl, C. ^{[1
]}

Chtcheglova, L. A.

Neundlinger, I.

Bozna, B. L.

Mayer, B. ^{[4
]}

Salio, M. ^{[2
]}

Shepherd, D. ^{[2
]}

Polzella, P. ^{[2
]}

Moertelmaier, M. ^{[1
]}

Kada, G. ^{[1
]}

Ebner, A. ^{[4
]}

Dieudonne, M. ^{[3
]}

Schuetz, G. J.

Cerundolo, V. ^{[2
]}

Kienberger, F. ^{[1
]}

Hinterdorfer, P. ^{[4
]}

机构：

[1] Agilent Technol Austria GmbH, A-4040 Linz, Austria

[2] Univ Oxford, Weatherall Inst Mol Med, Nuffield Dept Med, Canc Res UK Tumor Immunol Grp, Oxford OX3 9DS, England

[3] Agilent Technol Belgium, B-3110 Rotselaar, AN, Belgium

[4] Univ Linz, Inst Biophys, Christian Doppler Lab Nanoscop Methods Biophys, A-4040 Linz, Austria

来源：

NANOTECHNOLOGY | 2010年 / 21卷 / 11期

关键词：

ATOMIC-FORCE MICROSCOPY; VASCULAR ENDOTHELIAL-CELLS; AFM; ACTIVATION; STRATEGIES; ANTIBODIES;

D O I：

10.1088/0957-4484/21/11/115504

中图分类号：

TB3 [工程材料学];

学科分类号：

0805 ; 080502 ;

摘要：

The combination of fluorescence microscopy and atomic force microscopy has a great potential in single-molecule-detection applications, overcoming many of the limitations coming from each individual technique. Here we present a new platform of combined fluorescence and simultaneous topography and recognition imaging (TREC) for improved localization of cellular receptors. Green fluorescent protein (GFP) labeled human sodium-glucose cotransporter (hSGLT1) expressed Chinese Hamster Ovary (CHO) cells and endothelial cells (MyEnd) from mouse myocardium stained with phalloidin-rhodamine were used as cell systems to study AFM topography and fluorescence microscopy on the same surface area. Topographical AFM images revealed membrane features such as lamellipodia, cytoskeleton fibers, F-actin filaments and small globular structures with heights ranging from 20 to 30 nm. Combined fluorescence and TREC imaging was applied to detect density, distribution and localization of YFP-labeled CD1d molecules on alpha-galactosylceramide (alpha GalCer)-loaded THP1 cells. While the expression level, distribution and localization of CD1d molecules on THP1 cells were detected with fluorescence microscopy, the nanoscale distribution of binding sites was investigated with molecular recognition imaging by using a chemically modified AFM tip. Using TREC on the inverted light microscope, the recognition sites of cell receptors were detected in recognition images with domain sizes ranging from similar to 25 to similar to 160 nm, with the smaller domains corresponding to a single CD1d molecule.

引用

页数：7

共 28 条

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