Re-scan confocal microscopy (RCM) improves the resolution of confocal microscopy and increases the sensitivity
| Autor: | Ron A. Hoebe, Sjoerd Stallinga, Giulia M. R. De Luca, Ronald M. P. Breedijk, Erik M. M. Manders |
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| Přispěvatelé: | Cell Biology and Histology, Other Research, Medical Biology, Molecular Cytology (SILS, FNWI), Systems Biology |
| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Microscope Chemistry business.industry Confocal Resolution (electron density) Scanning confocal electron microscopy 01 natural sciences Atomic and Molecular Physics and Optics law.invention 010309 optics 03 medical and health sciences 030104 developmental biology Optics Confocal microscopy law 0103 physical sciences General Materials Science Pinhole (optics) Sensitivity (control systems) 4Pi microscope business Instrumentation Spectroscopy |
| Zdroj: | METHODS AND APPLICATIONS IN FLUORESCENCE, 5(1). IOP Publishing Ltd. Methods and applications in fluorescence, 5(1):015002. IOP Publishing Ltd. |
| ISSN: | 2050-6120 |
| Popis: | Re-scan confocal microscopy (RCM) is a new super-resolution technique based on a standard confocal microscope extended with a re-scan unit in the detection path that projects the emitted light onto a sensitive camera. In this paper the fundamental properties of RCM, lateral resolution, axial resolution and signal-to-noise ratio, are characterized and compared with properties of standard confocal microscopy. The results show that the lateral resolution of RCM is similar to 170 nm compared to similar to 240 nm of confocal microscopy for 488 nm excitation and 1.49 NA. As the theory predicts, this improved lateral resolution is independent of the pinhole diameter. In standard confocal microscopy, the same lateral resolution can only be achieved with an almost closed pinhole and, consequently, with a major loss of signal. We show that the sectioning capabilities of the standard confocal microscope are preserved in RCM and that the axial resolution of RCM is slightly better (similar to 15%) than the standard confocal microscope. Furthermore, the signal-to-noise ratio in RCM is a factor of 2 higher than in standard confocal microscopy, also due to the use of highly sensitive modern cameras. In case the pinhole of a confocal microscope is adjusted in such way that the lateral resolution is comparable to that of RCM, the signal-to-noise ratio in RCM is 4 times higher than standard confocal microscopy. Therefore, RCM offers a good alternative to standard confocal microscopy for higher lateral resolution with the main advantage of strongly improved sensitivity |
| Databáze: | OpenAIRE |
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